Morphic forms of trilaciclib and methods of manufacture thereof

ABSTRACT

An advantageous isolated morphic form of trilaciclib which is 2′-((5-(4-methylpiperazin-1-yl)pyridin-2-yl)amino)-7′,8′-dihydro-6′H-spiro[cyclohexane-1,9′-pyrazino[1′,2′:1, 5]pyrrolo[2,3 -d]pyrimidin]-6′-one, for example in the form of a di-hydrochloride salt or a dihydrochloride, dihydrate.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a continuation of U.S. patent application Ser. No.17/097,854, filed Nov. 13, 2020, which claims priority to U.S.Provisional Application 63/039,372, filed Jun. 15, 2020. The entirety ofeach of these applications is incorporated herein by reference.

FIELD OF THE INVENTION

This invention provides advantageous isolated morphic forms oftrilaciclib which is2′-((5-(4-methylpiperazin-1-yl)pyridin-2-yl)amino)-7′,8′-dihydro-6′H-spiro[cyclohexane-1,9′-pyrazino[1′,2′:1,5]pyrrolo[2,3-d]pyrimidin]-6′-one,for example in the form of a free base, a di-hydrochloride salt, or adihydrochloride, dihydrate.

BACKGROUND

U.S. Pat. Nos. 8,598,186; 8,598,197; 9,957,276 10,189,849; and10,189,850 and corresponding WO 2012/061156 assigned to G1 Therapeutics,Inc. describe a class of N-(heteroaryl)-pyrrolo[3,2-d]pyrimidin-2-aminecyclin dependent kinase inhibitors including2′-((5-(4-methylpiperazin-1-yl)pyridin-2-yl)amino)-7′,8′-dihydro-6′H-spiro[cyclohexane-1,9′-pyrazino[1′,2′:1,5]pyrrolo[2,3-d]pyrimidin]-6′-one(Compound 1) with the formula

Synthetic procedures to make Compound 1 are described in US 2019/135820and WO 2020/041770 both of which are assigned to G1 Therapeutics, Inc.

Compound 1 is referred to as “G1T28” or “trilaciclib.” It transientlyarrests normal cells to prevent chemotherapy-induced myelosuppressionand improve anti-tumor efficacy. It can be used, for example, inpatients with small cell lung cancer (SCLC) receiving topotecanchemotherapy, as well as in combination with etoposide and carboplatinin SCLC. It is also useful, for example, in combination withcarboplatin, etoposide, and atezolizumab (a PD-L1 inhibitor) in smallcell lung cancer (SCLC). Based on myelopreservation data in patientswith SCLC trilaciclib has been granted Breakthrough Therapy Designation(BTD) from the U.S. Food and Drug Administration (FDA) (see, e.g., Weisset al., “Myelopreservation with the CDK4/6 inhibitor trilaciclib inpatients with small-cell lung cancer receiving first-line chemotherapy:a phase Ib/randomized phase II trial,” Annals of Oncology 30:1613-1621(2019); Daniel et al., “Trilaciclib decreases myelosuppression inextensive-stage small cell lung cancer (ES-SCLC) patients receivingfirst-line chemotherapy plus atezolizumab,” ESMO 2019 Congress PosterAbstract #1742PD). Compound 1 is also in human clinical trials fortreatment of triple-negative breast cancer in combination with thestandard of care for neoadjuvant treatment of breast cancer.

SUMMARY

It has been discovered that Compound 1(2′-((5-(4-methylpiperazin-1-yl)pyridin-2-yl)amino)-7′,8′-dihydro-6′H-spiro[cyclohexane-1,9′-pyrazino[1′,2′:1,5]pyrrolo[2,3-d]pyrimidin]-6′-one)as a dihydrochloride can be prepared in a highly purified, advantageousmorphic form.

This highly purified, advantageous morphic form has been denoted“Pattern 1.” Pattern 1 is a highly crystalline form of Compound 1 as adihydrochloride. Pattern 1 exhibits superior stability when compared toother morphic forms. For example, in competitive slurry experiments (seeExample 10) Pattern 1 is the predominantly formed morphic structure ofCompound 1 as a dihydrochloride. Pattern 1 also has advantageousproperties in the manufacture of Compound 1 as a dihydrochloride. Forexample, Pattern 1 can be made on large scale via crystallization ofCompound 1 in a heated HCl solution. Pattern 1 can also be formed fromother morphic or amorphic forms of Compound 1 via recrystallization.

Compound 1 Pattern 1 can be in the form of a dihydrochloride, dihydrate.Pattern 1 typically, initially, forms a dihydrochloride, dihydrate andeven after being dried will eventually revert to a dihydrochloride,dihydrate when exposed to air. Regardless of water content Pattern 1 canmaintain its representative XRPD peaks as described in more detailbelow. For example, if the morphic form of Compound 1 is prepared asdescribed herein to form Pattern 1 and then dried, the representativeXRPD peaks will remain the same before and after drying.

Trilaciclib is used in certain embodiments as a myelopreservation agentto protect healthy cells, notably hematopoietic cells, duringchemotherapy. It is intended to be administered by intravenous injectionfor fast access to the blood stream just prior to administration ofchemotherapy, typically also by IV injection. However, a problem isevident that trilaciclib is not very soluble in water, and alsopractically insoluble in DMSO. Further, it becomes less soluble as thepH increases. Blood typically has a pH of 7.35-7.45, which is mildlybasic. Phosphate buffered saline typically has a pH of 7.2-7.4.Trilaciclib is not very soluble at this pH, and thus multiple problemscan occur. Most importantly, a significant amount of trilaciclib isrequired to achieve a therapeutic effect. A typical dose is 240 mg/m²and can be estimated to be about a 300 mg dose for a normal adult.Because of the lack of solubility in water, it would take a dilutesolution in a lot of fluid to provide this dosage of free basetrilaciclib, which would have to be given over a long period of time, incontradiction to how it should be delivered. The trilaciclib IV istypically administered over about 30 minutes (20-60 minutes), whichmeans that the drug has to be concentrated, not dilute, in the IV fluid.Also, when the free base trilaciclib is injected into blood, there is areal possibility of at least some of it falling out of solution becauseof the fact that blood is mildly basic. This can cause problems at thesite of injection caused by drug deposition and a loss of activity.

It has been discovered that an IV solution of trilaciclib for injectionto cancer patients to preserve healthy cells or for an antineoplasticcan be accomplished by administering it as a dihydrochloride or adihydrochloride, dihydrate. This enables the intended therapeutic effectbecause it can be delivered quickly in a concentrated form directly tothe blood stream. In a principal embodiment, the IV solution is preparedfrom a lyophilized powder of the dihydrochloride, dihydrate crystallinePattern 1.

In certain embodiments Compound 1 Pattern 1 is a dihydrochloride,dihydrate morphic form. In certain embodiments the invention providesthe composition of Compound 1 as a dihydrochloride, dihydrate. Incertain embodiments the invention provides the composition of Compound 1Pattern 1 as a dihydrochloride, dihydrate. In certain embodimentsCompound 1 Pattern 1 as a dihydrochloride, dihydrate can be treated forany condition that is responsive to trilaciclib, including formyelopreservation or as an antineoplastic agent.

Other morphic forms of Compound 1 as a dihydrochloride have also beenidentified. Pattern 2, and Pattern 3 are also highly crystalline formsof Compound 1 as a dihydrochloride. Pattern 4 is a crystalline solvateof Compound 1 as a dihydrochloride with acetonitrile. Pattern 5 is acrystalline form of Compound 1 as a dihydrochloride. Pattern 6 is ametastable form of Compound 1 as a dihydrochloride. Pattern 11 is acrystalline form of Compound 1 as a dihydrochloride, hemi-hydrate.

Morphic forms of Compound 1 as a free base have also been identified.Pattern 8 and Pattern 10 are crystalline forms of Compound 1 as a freebase. Pattern 9 is a crystalline form of Compound 1 as a free base whichmay be either a solvate or hydrate.

In certain embodiments Pattern 1 is the most stable morphic form ofCompound 1 as a dihydrochloride and formed by competitive slurryexperiments with either Pattern 2 or Pattern 3.

In one aspect of the present invention, isolated Compound 1 Pattern 1 isused in the manufacture of a lyophilized form that is then formulatedwith a suitable solvent such as phosphate buffered saline foradministration to a patient, for example, by intravenous delivery. Inanother embodiment it can be formulated into a parenteral dosage form.This dosage form can be used, for example, in subcutaneousadministration.

In another aspect of the present invention I Compound 1 Pattern 1 in theform of a dihydrochloride, dihydrate in a formulation is provided. Incertain embodiments the formulation comprises about 300 to 400 mg, about350-400 mg or more particularly about 373 mg of Compound 1 Pattern 1dihydrochloride, dihydrate. In a specific embodiment thepharmaceutically acceptable formulation includes about 373 mg ofCompound 1 Pattern 1 dihydrochloride, dihydrate (equivalent to about 300mg of trilaciclib free base), about 50-100 mg, about 60-80 mg, or moreparticularly about 76 mg of citric acid monohydrate, about 280-320 mg,for example 300 mg, of mannitol which may be provided as a lyophilizedform for later reconstitution.

Trilaciclib is typically administered to the patient in an intravenousformulation. However, the solid forms of trilaciclib can be importantbecause they may be used to achieve desired manufacturing purity and/orperformance, and may also be important to solid state shelf life priorto formulation, or in a solid dosage form for therapeutic delivery.

Given the therapeutic importance of trilaciclib to medical therapy forpatients suffering from a proliferative disorder such as a tumor orcancer, it is beneficial to provide this advantageous solid form of thecompound that contributes to the success or performance of the compoundduring manufacture, storage, formulation, and/or administration.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is an XRPD Diffractogram of the crystalline Pattern 1. Thecrystal structure was obtained using the method described in Example 1and the peaks are shown in Table 1. The x-axis is 2Theta measured indegrees and the y-axis is intensity measured in counts.

FIG. 2 is a PLM Imaging of the crystalline Pattern 1. The image wasobtained using the method described in Example 5. Pattern 1 materialappeared highly crystalline under PLM imaging with the non-polarizedmaterial shown on the left, and the polarized material shown on theright.

FIG. 3 is a thermogravimetric/differential thermal (TG/DTA) thermogramof crystalline Pattern 1. The TG/DTA thermogram of Pattern 1 wasobtained as described in Example 6. The initial mass loss of ca. 6% fromthe onset of heating related to the loss of surface moisture. Two masslosses of ca. 7% were observed from ca. 200° C. peaking at 218° C. and300° C. peaking at 343° C.

FIG. 4 is a Differential Scanning calorimetry (DSC) thermogram of thecrystalline Pattern 1. The DSC thermogram of crystalline Pattern 1 wasobtained as described in Example 7. DSC analysis showed 2 broadendotherms ca. 65° C. peaking at 130° C. and 245° C. peaking at 277° C.A sharp melting endotherm was observed from an onset of 330° C. peakingat 341° C.

FIG. 5. is a Dynamic Vapor Sorption (DVS) analysis showing the resultsfrom a moisture sorption experiment of Pattern 1. DVS analysis ofPattern 1 was obtained as described in Example 3. The material was foundto be stable and the XRPD analysis of a dried sample at the conclusionof the experiment confirmed Pattern 1. Pattern 1 adsorbed 3.00 wt % at40% RH (relative humidity) and 4.00 wt % at 90% RH. The x-axis isrelative humidity measured as a percent and the y-axis is weight ofwater of the material measured as a percent.

FIG. 6 is a Dynamic Vapor Sorption (DVS) Kinetic Plot of Pattern 1. DVSof Pattern 1 was obtained as described in Example 3. The sample wassubjected to a ramping profile from 40-90% relative humidity (RH) at 10%increments, maintaining the sample at each step until a stable weighthad been achieved (dm/dt 0.004%, minimum step length 30 minutes, maximumstep length 500 minutes) at 25° C. After completion of the sorptioncycle, the sample was dried using the same procedure to 0% RH and then asecond sorption cycle back to 40% RH. Two cycles were performed. Thematerial appeared hygroscopic by DVS with a mass increase of 4% between0% and 90% RH. During the desorption cycles the material dehydrates.

FIG. 7 is a comparison of XRPD diffractograms of Pattern 1 and Pattern 1post-DVS. The diffractogram of Pattern 1 was obtained as described inExample 1; and post-DVS of Pattern 1 was obtained as described inExample 11. The material appeared hygroscopic by DVS with a massincrease of 4% between 0% and 90% RH. During the desorption cycles thematerial dehydrates. XRPD analysis post-DVS showed no change in Pattern1. The x-axis is 2Theta measured in degrees and the y-axis is intensitymeasured in counts.

FIG. 8 is a comparison of XRPD diffractograms of Pattern 1 beforesolvent solubility studies and the Pattern 1 material that results fromvarious solvent solubility studies. The diffractograms were obtained asdescribed in Example 1 and the studies were conducted as described inExample 13. The x-axis is 2Theta measured in degrees and the y-axis isintensity measured in counts.

FIG. 9 is a comparison of XRPD diffractograms of Pattern 1 beforesolvent solubility studies and the Pattern 1 material that results fromvarious solvent solubility studies. The diffractograms were obtained asdescribed in Example 1 and the studies were conducted as described inExample 13. The x-axis is 2Theta measured in degrees and the y-axis isintensity measured in counts.

FIG. 10 is a comparison of XRPD diffractograms of Pattern 2 resultingfrom various solvent solubility studies. The diffractograms wereobtained as described in Example 1 and the studies were conducted asdescribed in Example 13. The x-axis is 2Theta measured in degrees andthe y-axis is intensity measured in counts.

FIG. 11 is a comparison of XRPD diffractograms of Pattern 2 resultingfrom various solvent solubility studies. The diffractograms wereobtained as described in Example 1 and the studies were conducted asdescribed in Example 13. The x-axis is 2Theta measured in degrees andthe y-axis is intensity measured in counts.

FIG. 12 is a comparison of XRPD diffractograms of Pattern 3 resultingfrom various solvent solubility studies. The diffractograms wereobtained as described in Example 1 and the studies were conducted asdescribed in Example 13. The x-axis is 2Theta measured in degrees andthe y-axis is intensity measured in counts.

FIG. 13 is a comparison of XRPD diffractograms of Pattern 1 and Pattern4 which resulted from the solubility study in MeCN. The diffractogramswere obtained as described in Example 1 and Example 4 and the studieswere conducted as described in Example 13. The x-axis is 2Theta measuredin degrees and the y-axis is intensity measured in counts.

FIG. 14 is a comparison of XRPD diffractograms Pattern 2 and Pattern 5which resulted from various solvent systems. The diffractograms wereobtained as described in Example 1 and the studies were conducted asdescribed in Example 13. The x-axis is 2Theta measured in degrees andthe y-axis is intensity measured in counts.

FIG. 15 is a comparison of XRPD diffractograms Pattern 2 and Pattern 6which resulted from various solvent systems. The diffractograms wereobtained as described in Example 1 and the studies were conducted asdescribed in Example 13. The x-axis is 2Theta measured in degrees andthe y-axis is intensity measured in counts.

FIG. 16 is a comparison of XRPD diffractograms of Pattern 1 before andafter maturation experiments in various solvents systems. Thediffractograms were obtained as described in Example 1 and the studieswere conducted as described in Example 21. The x-axis is 2Theta measuredin degrees and the y-axis is intensity measured in counts.

FIG. 17 is a comparison of XRPD diffractograms of Pattern 2 before andafter maturation experiments in various solvents systems. Thediffractograms were obtained as described in Example 1 and the studieswere conducted as described in Example 21. The x-axis is 2Theta measuredin degrees and the y-axis is intensity measured in counts.

FIG. 18 is a comparison of XRPD diffractograms of Pattern 2 before andafter maturation experiments in various solvents systems. Thediffractograms were obtained as described in Example 1 and the studieswere conducted as described in Example 21. The x-axis is 2Theta measuredin degrees and the y-axis is intensity measured in counts.

FIG. 19 is a comparison of XRPD diffractograms of Pattern 3 before andafter a maturation experiments in isopropyl alcohol and ethanol. Thediffractograms were obtained as described in Example 1 and the studieswere conducted as described in Example 21. The x-axis is 2Theta measuredin degrees and the y-axis is intensity measured in counts.

FIG. 20 is a comparison of XRPD diffractograms of Pattern 4 before andafter a maturation experiment in acetonitrile. The diffractograms wereobtained as described in Example 4 and the studies were conducted asdescribed in Example 21. The x-axis is 2Theta measured in degrees andthe y-axis is intensity measured in counts.

FIG. 21 is a comparison of XRPD diffractograms of Pattern 5 before andafter a maturation experiment in methyl isobutyl ketone. Thediffractograms were obtained as described in Example 1 and the studieswere conducted as described in Example 21. The x-axis is 2Theta measuredin degrees and the y-axis is intensity measured in counts.

FIG. 22 is a comparison of XRPD diffractograms resulting from maturationexperiments in various solvent systems. The diffractograms were obtainedas described in Example 1 and the studies were conducted as described inExample 21. The x-axis is 2Theta measured in degrees and the y-axis isintensity measured in counts.

FIG. 23 is a comparison of XRPD Pattern 1 diffractogram anddiffractograms resulting from maturation experiments in various solventsystems after drying. The diffractograms were obtained as described inExample 1 and the studies were conducted as described in Example 21. Thex-axis is 2Theta measured in degrees and the y-axis is intensitymeasured in counts.

FIG. 24 is a comparison of XRPD Pattern 1 diffractogram anddiffractograms resulting from maturation experiments in various solventsystems after drying. The diffractograms were obtained as described inExample 1 and the studies were conducted as described in Example 21. Thex-axis is 2Theta measured in degrees and the y-axis is intensitymeasured in counts.

FIG. 25 is a comparison of XRPD Pattern 2 diffractogram anddiffractograms resulting from maturation experiments in various solventsystems after drying. The diffractograms were obtained as described inExample 1 and the studies were conducted as described in Example 21. Thex-axis is 2Theta measured in degrees and the y-axis is intensitymeasured in counts.

FIG. 26 is a comparison of XRPD Pattern 3 diffractogram anddiffractograms resulting from maturation experiments in various solventsystems after drying. The diffractograms were obtained as described inExample 1 and the studies were conducted as described in Example 21. Thex-axis is 2Theta measured in degrees and the y-axis is intensitymeasured in counts.

FIG. 27 is a comparison of XRPD Pattern 3 diffractogram, Pattern 4diffractogram and diffractograms resulting from maturation experimentsin various solvent systems after drying. The diffractograms wereobtained as described in Example 1 and Example 4 and the studies wereconducted as described in Example 21. The x-axis is 2Theta measured indegrees and the y-axis is intensity measured in counts.

FIG. 28 is a comparison of XRPD Pattern 3 diffractogram, Pattern 6diffractogram and the diffractogram resulting from maturation in heptaneafter drying. The diffractograms were obtained as described in Example 1and the studies were conducted as described in Example 21. The x-axis is2Theta measured in degrees and the y-axis is intensity measured incounts.

FIG. 29 is a comparison of XRPD Pattern 5 and diffractograms resultingfrom maturation in various solvents after drying. The diffractogramswere obtained as described in Example 1 and the studies were conductedas described in Example 21. The x-axis is 2Theta measured in degrees andthe y-axis is intensity measured in counts.

FIG. 30 is a comparison of XRPD diffractogram of Pattern 1 and solidsthat were left after evaporation experiments. The diffractograms wereobtained as described in Example 1. The solids were obtained asdescribed in Example 13. Samples were left uncapped and placed in acupboard to allow evaporation. The x-axis is 2Theta measured in degreesand the y-axis is intensity measured in counts.

FIG. 31 is a comparison of XRPD diffractogram of Pattern 3 and solidsthat were left after evaporation experiments. The diffractograms wereobtained as described in Example 1. The solids were obtained asdescribed in Example 13. Samples were left uncapped and placed in acupboard to allow evaporation. The x-axis is 2Theta measured in degreesand the y-axis is intensity measured in counts.

FIG. 32 is a comparison of XRPD diffractogram of Pattern 3 and the solidthat resulted from the addition of methyl tent-butyl ether (t-BME) to2,2,2-trifluoroethanol. The diffractograms were obtained as described inExample 1. The solids were obtained as described in Example 13. Thex-axis is 2Theta measured in degrees and the y-axis is intensitymeasured in counts.

FIG. 33 is a thermogravimetric/differential thermal (TG/DTA) thermogramof Pattern 2 small scale material. The TG/DTA thermogram of Pattern 2was obtained as described in Example 6. The initial mass loss of ca. 3%from the onset of heating related to the loss of surface moisture. Twomass losses of ca. 7% were observed from ca. 222° C. peaking at 239° C.,318° C. peaking at 340° C., and 345° C. peaking at 325° C. The x-axis inTemp (° C.) and the y-axis is TG (%).

FIG. 34 is a thermogravimetric/differential thermal (TG/DTA) thermogramof Pattern 3 small scale material. The TG/DTA thermogram of Pattern 3was obtained as described in Example 6. The initial mass loss of ca.2.6% from the onset of heating related to the loss of surface moisture.Two mass losses of ca. 6.6% were observed from ca. 222° C. peaking at229° C., 311° C. peaking at 317° C., and 343° C. peaking at 345° C. Thex-axis in Temp (° C.) and the y-axis is TG (%).

FIG. 35 is a thermogravimetric/differential thermal (TG/DTA) thermogramof Pattern 5 small scale material. The TG/DTA thermogram of Pattern 5was obtained as described in Example 6. The initial mass loss of ca.3.4% from the onset of heating related to the loss of surface moisture.Three mass losses of ca. 7.3% were observed from ca. 206° C. peaking at216° C., 284° C. peaking at 321° C. and 343° C. peaking at 345° C. Thex-axis in Temp (° C.) and the y-axis is TG (%).

FIG. 36 is a comparison of XRPD patterns for small scale and scale-upmaterials of Pattern 2 and Pattern 6. When the Pattern 2 synthesis wasscaled it produced a mixture of Pattern 6 and Pattern 2 as shown in theXRPD diffractograms. The diffractograms were obtained as describedExample 1. The x-axis is 2Theta measured in degrees and the y-axis isintensity measured in counts.

FIG. 37 is a PLM image of the mixture of Pattern 2 and Pattern 6 thatresulted from the scaleup attempt. The PLM imaging of Pattern 2/6mixture was obtained as described in Example 5. Pattern 2/6 scale-upmaterial appeared highly crystalline under PLM imaging with thenon-polarized material shown on the left, and the polarized materialshown on the right.

FIG. 38 is a thermogravimetric/differential thermal (TG/DTA) thermogramof the mixture of Pattern 2 and Pattern 6 that resulted from the scaleupattempt. The initial mass loss of ca. 3.6% from the onset of heatingrelated to the loss of surface moisture. Two mass losses of ca. 6.3%were observed from ca. 211° C. and 310° C. peaking at 321° C. The x-axisin Temp (° C.) and the y-axis is TG (%).

FIG. 39 is a differential scanning calorimetry (DSC) thermogram of themixture of Pattern 2 and Pattern 6 that resulted from the scaleupattempt. The DSC thermogram of Pattern 2/6 mixture was obtained asdescribed in Example 7. DSC analysis showed 2 broad endotherms ca. 55.9°C. peaking at 131° C. and 267° C. peaking at 295° C. A sharp meltingendotherm was observed from an onset of 336° C.

FIG. 40 is a gravimetric vapor sorption (GVS) isotherm plot of themixture of Pattern 2 and Pattern 6 that resulted from the scaleupattempt. The GVS isotherm plot of Pattern 2/6 mixture was obtained asdescribed in Example 2. The mixture of pattern 2 and pattern 6 adsorbed4.00% wt % at 40% RH (relative humidity) and 7.00% wt % at 90% RH. Thex-axis is relative humidity measured as a percent and the y-axis isweight of water of the material measured as a percent.

FIG. 41 is a gravimetric vapor sorption (GVS) kinetic plot of themixture of Pattern 2 and Pattern 6 that resulted from the scaleupattempt. The GVS isotherm plot of Pattern 2/6 mixture was obtained asdescribed in Example 2. The sample was subjected to a ramping profilefrom 40-90% relative humidity (RH) at 10% increments, maintaining thesample at each step until a stable weight had been achieved (dm/dt0.004%, minimum step length 550 minutes, maximum step length 900minutes) at 25° C. After completion of the sorption cycle, the samplewas dried using the same procedure to 0% RH and then a second sorptioncycle back to 40% RH. Two cycles were performed. The material appearedhygroscopic by DVS with a mass increase of 8% between 0% and 90% RH.During the desorption cycles the material dehydrates.

FIG. 42 is a comparison of XRPD diffractograms of Pattern 1, Pattern 2scale up and Pattern 2 scale up post-DVS. The diffractograms wereobtained as described in Example 1. The x-axis is 2Theta measured indegrees and the y-axis is intensity measured in counts.

FIG. 43 is a comparison of XRPD diffractograms of Pattern 3 small scaleand Pattern 3 scaled in dry and damp form. The diffractogram wereobtained as described in Example 1. The x-axis is 2Theta measured indegrees and the y-axis is intensity measured in counts.

FIG. 44 is a PLM image of the Pattern 3 scale-up material. Pattern 3scale-up material appeared highly crystalline under PLM imaging asdescribed in Example 5. The non-polarized material shown on the left,and the polarized material shown on the right.

FIG. 45 is a TG/DTA thermogram of the Pattern 3 scale up material. TheTG/DTA thermogram of Pattern 3 scale-up was obtained as described inExample 6. The TG/DTA shows two peaks at 199° C. and 345° C. The initialmass loss of ca. 8% from the onset of heating related to the loss ofsurface moisture. Two mass losses of ca. 7% were observed from ca. 199°C. and 343° C. peaking at 345° C. The x-axis in Temp (° C.) and they-axis is TG (%).

FIG. 46 is a differential scanning calorimetry (DSC) thermogram of thePattern 3 scale-up material. The DSC thermogram of Pattern 3 wasobtained as described in Example 7. DSC analysis showed 3 broadendotherms ca. 168° C. peaking at 189° C., 251° C. peaking at 264° C.and 272° C. peaking at 296° C. A sharp melting endotherm was observedfrom an onset of 337° C. peaking at 343° C.

FIG. 47 is a DVS isotherm plot of the Pattern 3 scale-up material. TheDVS isotherm plot of Pattern 3 was obtained as described in Example 3.The material appeared hygroscopic by DVS with a mass increase of 4%between 0% and 90% RH. During the desorption cycles the materialdehydrates.

FIG. 48 is a DVS kinetic plot of the Pattern 3 scale-up material. TheDVS kinetic plot of Pattern 3 was obtained as described in Example 3.The sample was subjected to a ramping profile from 40-90% relativehumidity (RH) at 10% increments, maintaining the sample at each stepuntil a stable weight had been achieved (change in mass 0.004%, minimumstep length 650 minutes, maximum step length 1050 minutes) at 25° C.After completion of the sorption cycle, the sample was dried using thesame procedure to 0% RH and then a second sorption cycle back to 40% RH.Two cycles were performed. The material appeared hygroscopic by DVS witha mass increase of 4% between 0% and 90% RH. During the desorptioncycles the material dehydrates.

FIG. 49 is a comparison of XRPD diffractograms of the Pattern 3 scale-upmaterial before and after DVS. The diffractograms were obtained asdescribed in Example 1. The x-axis is 2Theta measured in degrees and they-axis is intensity measured in counts.

FIG. 50 is a comparison of the XRPD diffractograms of Pattern 5 fromsmall scale and scale up experiments. The diffractogram were obtained asdescribed in Example 1. The x-axis is 2Theta measured in degrees and they-axis is intensity measured in counts.

FIG. 51 is a comparison of the XRPD diffractograms of Pattern 5 fromsmall scale and scale up experiments after repeating the process. Thediffractogram were obtained as described in Example 1. The x-axis is2Theta measured in degrees and the y-axis is intensity measured incounts.

FIG. 52 is a comparison of XRPD diffractograms of Pattern 2 in variousweeklong stability studies. The diffractograms were obtained asdescribed in Example 1. The stability study compares Pattern 2, andPattern 2 after 1 week at ambient temperature and at 80° C.,respectively. The x-axis is 2Theta measured in degrees and the y-axis isintensity measured in counts. Repeat Scale Up.

FIG. 53 is a comparison of XRPD diffractograms of Pattern 2 in variousweeklong stability studies. The diffractograms were obtained asdescribed in Example 1. The stability study compares Pattern 2, Pattern2 scale-up and Pattern 2 at 40° C. The x-axis is 2Theta measured indegrees and the y-axis is intensity measured in counts. Repeat Scale Up.

FIG. 54 is a comparison of XRPD diffractograms of Pattern 3 in variousweeklong stability studies. The diffractograms were obtained asdescribed in Example 1. The stability study compares Pattern 3, andPattern 3 after 1 week at ambient temperature, 40° C. and at 80° C.,respectively. The x-axis is 2Theta measured in degrees and the y-axis isintensity measured in counts.

FIG. 55 is a comparison of XRPD diffractograms resulting fromcompetitive slurry experiments of Pattern 1 mixed with Pattern 2 andPattern 3. The diffractograms were obtained as described in Example 1after the procedure form Example 11 was pursued. The x-axis is 2Thetameasured in degrees and the y-axis is intensity measured in counts.

FIG. 56 is a comparison of XRPD diffractograms resulting fromcompetitive slurry experiments of Pattern 1 mixed with Pattern 2 andPattern 3. The diffractograms were obtained as described in Example 1after the procedure form Example 11 was pursued. The x-axis is 2Thetameasured in degrees and the y-axis is intensity measured in counts.

FIG. 57 is a comparison of XRPD diffractograms resulting fromcompetitive slurry experiments of Pattern 1 mixed with Pattern 2 andPattern 3. The diffractograms were obtained as described in Example 1after the procedure form Example 11 was pursued. The x-axis is 2Thetameasured in degrees and the y-axis is intensity measured in counts.

FIG. 58 is a comparison of XRPD diffractograms resulting fromcompetitive slurry experiments of Pattern 1 mixed with Pattern 2 andPattern 3. The diffractograms were obtained as described in Example 1after the procedure form Example 11 was pursued. The x-axis is 2Thetameasured in degrees and the y-axis is intensity measured in counts.

FIG. 59 is a comparison of XRPD diffractograms resulting fromcompetitive slurry experiments of Pattern 1 mixed with Pattern 2 andPattern 3. The diffractograms were obtained as described in Example 1after the procedure form Example 11 was pursued. The x-axis is 2Thetameasured in degrees and the y-axis is intensity measured in counts.

FIG. 60 is a comparison of XRPD diffractograms of Pattern 7 prior to andafter thermodynamic solubility experiments. The thermodynamic solubilityexperiment is described in Example 22. The diffractograms were obtainedusing the procedure from Example 1. The x-axis is 2Theta measured indegrees and the y-axis is intensity measured in counts.

FIG. 61 is an XRPD diffractogram of Pattern 7. The diffractogram ofPattern 7 was obtained as described in Example 1. The x-axis is 2Thetameasured in degrees and the y-axis is intensity measured in counts.

FIG. 62 is a PLM image of Pattern 7 at 200× Magnification. The PLM imageof Pattern 7 was obtained as described in Example 5. The birefringentsample under polarized light is shown on the bottom and theNon-Polarized Light of the sample is shown on the top.

FIG. 63 is a TG/DTA thermogram of Pattern 7. The TG/DTA thermogram ofPattern 7 was obtained as described in Example 6. The thermogram shows a6.2% weight loss between 150 and 340° C. and sample decomposition above350° C. The thermogram also shows that the sample melted noted withonset at 340° C. and a peak at 345° C. The x-axis is Temp measured indegrees Celsius and the y-axis is intensity measured in DTAuV on theleft and TG % on the right.

FIG. 64 is a DSC first heat thermogram of Pattern 7. The DSC thermogramof Pattern 7 was obtained as described in Example 7. The thermogramshows the sample melting event at 335° C. and a peak at 338° C.(consistent with TG/DTA data) during first heat. The x-axis is Tempmeasured in degrees Celsius and the y-axis is intensity measured in DSCmW.

FIG. 65 is a DSC cool thermogram of Pattern 7. The DSC thermogram ofPattern 7 was obtained as described in Example 7. The thermogram showsno significant thermal events in the cool cycle. The x-axis is Tempmeasured in degrees Celsius and the y-axis is intensity measured in DSCmW.

FIG. 66 is a DSC second heat thermogram of the crystalline Pattern 7.The DSC thermogram of Pattern 7 was obtained as described in Example 7.The second heat contained the melting event with onset 333° C. and apeak at 339° C. Possible incomplete melting during the initial heatcycle. The x-axis is Temp measured in degrees Celsius and the y-axis isintensity measured in DSC mW.

FIG. 67 is a comparison of XRPD diffractograms from pH solubilitystudies of Pattern 7 at various pHs. The material resulting from pH 4was identified as Pattern 8. The diffractograms were obtained asdescribed in Example 1 and the pH experiment was as described in Example16. The x-axis is 2Theta measured in degrees and the y-axis is intensitymeasured in counts.

FIG. 68 is a comparison of XRPD diffractograms from pH solubilitystudies of Pattern 7 at various pHs. The diffractogram of Pattern 7 wasobtained as described in Example 1 and Example 16. The x-axis is 2Thetameasured in degrees and the y-axis is intensity measured in counts.

FIG. 69 is a comparison of XRPD diffractograms from pH solubilitystudies of Pattern 7 at various pH solvent systems. The diffractogram ofPattern 7 was obtained as described in Example 1 and Example 16. Thex-axis is 2Theta measured in degrees and the y-axis is intensitymeasured in counts.

FIG. 70 is a comparison of XRPD diffractograms that resulted fromcrystallization set 1. The crystallizations were conducted as describedin Example 17. The comparison shows the Pattern 7 and forms resultingfrom various concentrations in various pH solvent systems. The x-axis is2Theta measured in degrees and the y-axis is intensity measured incounts.

FIG. 71 is a comparison of XRPD diffractograms that resulted fromcrystallization sets 2 and 3. The crystallizations were conducted asdescribed in Example 18 and Example 19. The comparison shows the Pattern7 and forms resulting from various concentrations in various pH solventsystems. The x-axis is 2Theta measured in degrees and the y-axis isintensity measured in counts.

FIG. 72 is a comparison of XRPD diffractograms that resulted fromcrystallization sets 2 and 3. The crystallizations were conducted asdescribed in Example 18 and Example 19. The comparison shows the Pattern7 and forms resulting from various concentrations in various pH solventsystems. The x-axis is 2Theta measured in degrees and the y-axis isintensity measured in counts.

FIG. 73 is a comparison of XRPD diffractograms that resulted fromcrystallization set 4. The crystallizations were conducted as describedin Example 20. The comparison shows the Pattern 7 and forms resultingfrom various concentrations in various pH solvent systems. The x-axis is2Theta measured in degrees and the y-axis is intensity measured incounts.

FIG. 74 is a PLM image corresponding to the material obtained fromexperiment 1 from Example 23. The polarized sample is shown in thebottom row and the non-polarized light sample is shown in the top row.

FIG. 75 is a PLM image corresponding to the material obtained fromexperiment 2 from Example 23. The PLM image was obtained as described inExample 5. The polarized sample is shown on the right and thenon-polarized light sample is shown on the left.

FIG. 76 is a PLM image corresponding to the material obtained fromexperiment 3 from Example 23. The PLM image was obtained as described inExample 5. The polarized sample is shown on the right and thenon-polarized light sample is shown on the left.

FIG. 77 is a PLM image corresponding to the material obtained fromexperiment 4 from Example 23. The PLM image was obtained as described inExample 5. The polarized sample is shown in the bottom row and thenon-polarized light sample is shown in the top row.

FIG. 78 is a PLM image corresponding to the material obtained fromexperiment 5 from Example 23. The PLM image was obtained as described inExample 5. The polarized sample is shown in the bottom row and theNon-Polarized Light sample is shown in the top row.

FIG. 79 is a PLM image corresponding to the material obtained fromexperiment 7 from Example 23. The PLM image was obtained as described inExample 5. The polarized sample is shown on the right and thenon-polarized Light sample is shown on the left.

FIG. 80 is a PLM image corresponding to the material obtained fromexperiment 6 from Example 23. The PLM image was obtained as described inExample 5. The polarized sample is shown in the bottom row and theNon-Polarized Light sample is shown in the top row.

FIG. 81 is a PLM image corresponding to the material obtained fromexperiment 8 from Example 23. The polarized sample is shown on the rightand the non-polarized Light sample is shown on the left.

FIG. 82 is a PLM image corresponding to the material obtained fromexperiment 9 from Example 23. The PLM image was obtained as described inExample 5. The polarized sample is shown on the right and thenon-polarized Light sample is shown on the left.

FIG. 83 is a PLM image corresponding to the material obtained fromexperiment 10 from Example 23. The polarized sample is shown in thebottom row and the Non-Polarized Light sample is shown in the top row.

FIG. 84 is a PLM image corresponding to the material obtained fromexperiment 11 from Example 23. The polarized sample is shown on theright and the non-polarized Light sample is shown on the left.

FIG. 85 is a PLM image corresponding to the material obtained fromexperiment 12 from Example 23. This material was identified as Pattern9. The polarized sample is shown in the bottom row and the Non-PolarizedLight sample is shown in the top row.

FIG. 86 is a PLM Images of the crystalline Pattern 10 at 200×Magnification. The PLM imaging of crystalline Pattern 7 was obtained asdescribed in Example 5. The polarized sample is shown in the bottom rowand the Non-Polarized Light sample is shown in the top row.

FIG. 87 is a TG/DTA thermogram of Pattern 9. The TG/DTA thermogram ofPattern 9 was obtained as described in Example 6. Pattern 9 exhibited alarge weight loss of 24.3% from the onset of heating up to 150° C.Pattern 9 exhibited a second mass loss of 15 wt. % between 75° C. and154° C. The DT trace identified two endothermic events associated withthe mass losses at the first shown onset at 37° C. peaking at 66.2° C.;the second onset at 122° C. peaking at 128° C.; and the third and finalendothermic event in the DT trace (sample melt) was observed with anonset of 326° C. and a peak at 330° C. The x-axis is Temp measured indegrees Celsius and the y-axis is intensity measured in DTAuV on theleft and TG % on the right.

FIG. 88 is a TG/DTA thermogram of Pattern 10. The TG/DTA thermogram wasobtained as described in Example 6. The thermogram shows showed minimalweight loss of 0.1% weight loss between 150 and 340° C. from onset ofheating. The sample showed degradation above 320° C. The thermogram alsoshows two peaks, with the first onset at 199 and peaking at 206; and thesecond onset at 327° C. peaking at 331° C. The x-axis is Temp measuredin degrees Celsius and the y-axis is intensity measured in DTAuV on theleft and TG % on the right.

FIG. 89 is an XRPD Diffractogram of the crystalline Pattern 8. Thecrystal structure was obtained using the method described in Example 1and the peaks are shown in Table 1. The x-axis is 2Theta measured indegrees and the y-axis is intensity measured in counts.

FIG. 90 is the crystal structure of Pattern 11. The crystal structurewas solved as described in Example 25. Non-hydrogen atoms are shown withthermal ellipsoids set at 50% probability level.

FIG. 91 is the crystal structure of Pattern 11 with hydrogen bondingdepicted. The crystal structure was solved as described in Example 25.Non-hydrogen atoms are shown with thermal ellipsoids set at 50%probability level.

FIG. 92 is the crystal structure of Pattern 11 with the view set to theunit cell a axis. The crystal structure was solved as described inExample 25. Non-hydrogen atoms are shown with thermal ellipsoids set at50% probability level.

FIG. 93 is the crystal structure of Pattern 11 with the view set to theunit cell b axis. The crystal structure was solved as described inExample 25. Non-hydrogen atoms are shown with thermal ellipsoids set at50% probability level.

FIG. 94 is the crystal structure of Pattern 11 with the view set to theunit cell c axis. The crystal structure was solved as described inExample 25. Non-hydrogen atoms are shown with thermal ellipsoids set at50% probability level.

FIG. 95 is the crystal structure of Pattern 11 with the view set to thevoid view of the a axis. The crystal structure was solved as describedin Example 25. Non-hydrogen atoms are shown with thermal ellipsoids setat 50% probability level.

FIG. 96 is a comparison of the simulated XRPD pattern of Pattern 11 andthe XRPD pattern of Pattern 1. The x-axis is 2Theta measured in degreesand the y-axis is intensity measured in counts.

FIG. 97 is the crystal structure of Pattern 1 resulting from the x-raycrystallography discussed in Example 27.

FIG. 98 is the crystal structure of Pattern 1 resulting from the x-raycrystallography discussed in Example 27 where the hydrogen bonds areshown explicitly.

FIG. 99 is an overlay of the experimentally determined XRPDdiffractogram of Pattern 1 with the simulated diffractogram thatcorresponds to the crystal structure shown in FIG. 97. The differencesbetween these two lines is shown on the bottom line. As the bottom lineis nearly flat the simulation strongly suggests that the crystalstructure corresponds to the XRPD pattern.

DETAILED DESCRIPTION OF THE INVENTION

It cannot be predicted in advance whether a compound exists in more thanone solid form or with which salt or solvate it exists or what thevarious properties of any solid form might be if one or more does exist,or whether the properties are advantageous for a therapeutic dosageform. As one example, the drug ritonavir is active in one morphic formand inactive in another form, and the inactive form is the more stable.

Trilaciclib is used in certain embodiments as a myelopreservation agentto protect healthy cells, notably hematopoietic cells, duringchemotherapy. It is intended to be administered by intravenous injectionfor fast access to the blood stream just prior to administration ofchemotherapy, typically also by IV injection. However, a problem isevident that trilaciclib is not very soluble in water, and alsopractically insoluble in DMSO. Further, it becomes less soluble as thepH increases. Blood typically has a pH of 7.35-7.45, which is mildlybasic. Phosphate buffered saline typically has a pH of 7.2-7.4.Trilaciclib is not very soluble at this pH, and thus multiple problemscan occur. Most importantly, a significant amount of trilaciclib isrequired to achieve a therapeutic effect. A typical dose is 240 mg/m²and can be estimated to be about a 300 mg dose for a normal adult.Because of the lack of solubility in water, it would take a dilutesolution in a lot of fluid to provide this dosage of free basetrilaciclib, which would have to be given over a long period of time, incontradiction to how it should be delivered. The trilaciclib IV istypically administered over about 30 minutes (20-60 minutes), whichmeans that the drug has to be concentrated, not dilute, in the IV fluid.Also, when the free base trilaciclib is injected into blood, there is areal possibility of at least some of it falling out of solution becauseof the fact that blood is mildly basic. This can cause problems at thesite of injection caused by drug deposition and a loss of activity.

It has been discovered that an IV solution of trilaciclib for injectionto cancer patients to preserve healthy cells or for an antineoplastictreatment can be accomplished by administering it as a dihydrochlorideor a dihydrochloride, dihydrate. This enables the intended therapeuticeffect because it can be delivered quickly in a concentrated formdirectly to the blood stream. In a principal embodiment, the IV solutionis prepared from a lyophilized powder of the dihydrochloride, dihydratecrystalline Pattern 1.

Pattern 1, Pattern 2, and Pattern 3 are highly crystalline forms ofCompound 1 as a dihydrochloride. Pattern 1 is a useful dihydrochloride,dihydrate form of Compound 1. Pattern 4 is a crystalline solvate ofCompound 1 as a dihydrochloride with acetonitrile. Pattern 5 is acrystalline form of Compound 1 as a dihydrochloride. Pattern 6 is ametastable form of Compound 1 as a dihydrochloride. Pattern 7 is ahighly crystalline form of Compound 1 as a free base. Pattern 8 andPattern 10 are crystalline forms of Compound 1 as a free base. Pattern 9is a crystalline form of Compound 1 as a free base which may be either asolvate or hydrate. Pattern 11 is a crystalline form of Compound 1 as adihydrochloride, hemi-hydrate.

In certain embodiments Pattern 1 dihydrochloride, dihydrate is the moststable morphic form of Compound 1 as a dihydrochloride and can be formedby competitive slurry experiments with either Pattern 2 or Pattern 3. Incertain embodiments Pattern 1 regardless of its water content is themost stable morphic form of Compound 1 as a dihydrochloride and can beformed by competitive slurry experiments with either Pattern 2 orPattern 3. In certain embodiments Pattern 7 is the most stable morphicform of Compound 1 as a free base.

Pattern 1 dihydrochloride, dihydrate has multiple therapeutic advantagesover the other morphic forms of Compound 1. For example, Pattern 1dihydrochloride, dihydrate is more easily used in an intravenoussolution. Additionally, higher concentrations of Pattern 1dihydrochloride, dihydrate can be achieved in water and/or DMSO.

Pattern 1 dihydrochloride, dihydrate has multiple manufacturingadvantages over the other morphic forms of Compound 1. For example,Pattern 1 dihydrochloride, dihydrate typically has increased shelfstability, thermodynamic stability, and/or solubility in water thanother morphic forms of Compound 1. In other embodiments the manufactureof Pattern 1 dihydrochloride, dihydrate is reproducibly more scalablethan the manufacture of other morphic forms of Compound 1.

In one aspect of the present invention, isolated Pattern 1, optionallyas a dihydrate, is used in the manufacture of a lyophilized form that isthen formulated with a suitable solvent such as phosphate bufferedsaline for administration to a patient, for example, by intravenousdelivery. In an alternative embodiment it can be formulated into aparenteral dosage form. This dosage form can be used, for example, insubcutaneous administration.

In one aspect of the present invention, isolated Pattern 2 is used inthe manufacture of a lyophilized form that is then formulated with asuitable solvent such as phosphate buffered saline for administration toa patient, for example, by intravenous delivery. In an alternativeembodiment it can be formulated into a parenteral dosage form. Thisdosage form can be used, for example, in subcutaneous administration.

In one aspect of the present invention, isolated Pattern 3 is used inthe manufacture of a lyophilized form that is then formulated with asuitable solvent such as phosphate buffered saline for administration toa patient, for example, by intravenous delivery. In an alternativeembodiment it can be formulated into a parenteral dosage form. Thisdosage form can be used, for example, in subcutaneous administration.

In one aspect of the present invention, isolated Pattern 7 is used inthe manufacture of a lyophilized form that is then formulated with asuitable solvent such as phosphate buffered saline for administration toa patient, for example, by intravenous delivery. In an alternativeembodiment it can be formulated into a parenteral dosage form. Thisdosage form can be used, for example, in subcutaneous administration.

Pattern 1

An isolated morphic form of Compound 1 as a dihydrochloride that hasbeen designated Pattern 1 is provided in this invention.

Compound 1 Pattern 1 can be in the form of a dihydrochloride, dihydrate.Pattern 1 is typically, initially, in the form of a dihydrochloride,dihydrate after being formed and even after being dried will eventuallyrevert to a dihydrochloride, dihydrate when exposed to air. Regardlessof water content Pattern 1 can maintain its representative XRPD peaks asdescribed in more detail below. For example, if the morphic form ofCompound 1 is prepared as described herein to form Pattern 1 and thendried, the representative XRPD peaks will remain the same before andafter drying.

In certain embodiments Compound 1 Pattern 1 is a dihydrochloride,dihydrate morphic form. In certain embodiments the invention providesthe composition of Compound 1 as a dihydrochloride, dihydrate. Incertain embodiments the invention provides the composition of Compound 1Pattern 1 as a dihydrochloride salt dihydrate.

In certain embodiments Compound 1 Pattern 1 dihydrochloride, dihydrateis more stable than other hydrate forms of Compound 1. In anotherembodiment Compound 1 Pattern 1 dihydrochloride, dihydrate is morestable than non-hydrate forms of Compound 1. Compound 1 Pattern 1 as adihydrochloride, dihydrate can be advantageous to the anhydrous,monohydrate, or even trihydrate versions of Compound 1 Pattern 1. Forexample, the dihydrate version of Compound 1 Pattern 1 dihydrochloridehas increased shelf stability, thermodynamic stability, and/orsolubility in water than the anhydrous, monohydrate, or even trihydrateversions of Compound 1 Pattern 1. In other embodiments the manufactureof Pattern 1 dihydrochloride, dihydrate is cheaper, faster, and/or morescalable than the manufacture of the anhydrous, monohydrate, or eventrihydrate versions Compound 1 Pattern 1.

In certain embodiments, Compound 1 Pattern 1, optionally as a dihydrate,is characterized by an XRPD pattern that is substantially similar tothat set forth in FIG. 1. In one embodiment, Compound 1 Pattern 1optionally as a dihydrate, is characterized by an XRPD patterncomprising at least three 2theta values selected from about 9.6±0.2°,about 21.3±0.2°, about 19.8±0.2°, about 12.2±0.2°, about 24.0±0.2°,about 26.1±0.2°, about 19.3±0.2°, about 17.6±0.2°, and about 28.6±0.2°.In one embodiment, Compound 1 Pattern 1, optionally as a dihydrate, ischaracterized by an XRPD pattern comprising a peak with a 2theta valueof about 9.6±0.2°.

In certain embodiments, Compound 1 Pattern 1, optionally as a dihydrate,is characterized as having a 6%±3% weight loss on the onset of heatingthe sample. In certain embodiments, Compound 1 Pattern 1, optionally asa dihydrate, is characterized as having a 7%±3% weight loss betweenabout 200° C. and about 230° C. In certain embodiments, Compound 1Pattern 1 is characterized as having a second 7%±3% weight loss betweenabout 300° C. and about 380° C.

Compound 1 Pattern 1 can be prepared using selective crystallization.The method can be carried out by treating a solution comprising asuitable solvent(s) and Compound 1 as a dihydrochloride optionally inthe presence of one or more seeds comprising Compound 1 Pattern 1 toconditions that provide for the crystallization of Compound 1 Pattern 1.If the solvent used is water or contains water then the resultingCompound 1 Pattern 1 is typically a dihydrochloride, dihydrate. Theselective crystallization can be carried out in any suitable solvent.For example, it can be carried out in an aprotic solvent or a mixturethereof. Examples of aprotic solvents that can be used includetetrahydrofuran, dichloromethane, nitromethane, and dioxane. In anotherembodiment Compound 1 Pattern 1 is prepared from a protic solvent.Examples of protic solvents that can be used include water, methanol,and ethanol. In certain embodiments Compound 1 Pattern 1 is preparedfrom a mixture of an aprotic solvent and a protic solvent, such as forexample a mixture of acetonitrile and water.

The selective crystallization can be carried out at, for example, atemperature in the range of about 20° C. to about 65° C. In anotherembodiment the selective crystallization can be carried out at, forexample, a temperature in the range of about 35° C. to about 45° C. orat about 40° C.

In one embodiment, Compound 1 Pattern 1, optionally as a dihydrate, isproduced by crystallization or recrystallization in an acidic solution.For example Compound 1 Pattern 1 can be produced in an aqueoushydrochloric acid solution.

In one embodiment Compound 1 Pattern 1 is produced by heating Compound 1to about 80° C. and agitating for at least 30 minutes. The resultingsolution is filtered into a reactor pre-heated to about 80° C., and thereaction mixture is cooled to about 70° C. Preheated (about 70° C.)purified water is added and the reaction mixture is cooled to below 20°C. with agitation. Compound 1 Pattern 1 can be isolated by filtration,washed with purified water and acetone, and dried under vacuum at anelevated temperature.

In certain embodiments Compound 1 Pattern 1, optionally as a dihydrate,is sieved using a mill. In certain embodiments the sieved Compound 1Pattern 1 material is formulated for parenteral administration to apatient. In certain embodiments, Compound 1 Pattern 1, optionally as adihydrate, is characterized by an XRPD pattern comprising at least 2peaks selected from about 9.6±0.2°, about 12.2±0.2°, about 19.8±0.2°,about 21.2±0.2°, about 23.9±0.2° and about 26.1±0.2°.

In certain embodiments, Compound 1 Pattern 1, optionally as a dihydrate,is characterized by an XRPD pattern comprising at least 3 peaks selectedfrom about 9.6±0.2°, about 12.2±0.2°, about 19.8±0.2°, about 21.2±0.2°,about 23.9±0.2° and about 26.1±0.2°.

In certain embodiments, Compound 1 Pattern 1, optionally as a dihydrate,is characterized by an XRPD pattern comprising at least 4 peaks selectedfrom about 9.6±0.2°, about 12.2±0.2°, about 19.8±0.2°, about 21.2±0.2°,about 23.9±0.2° and about 26.1±0.2°.

In certain embodiments, Compound 1 Pattern 1, optionally as a dihydrate,is characterized by an XRPD pattern comprising at least 5 peaks selectedfrom about 9.6±0.2°, about 12.2±0.2°, about 19.8±0.2°, about 21.2±0.2°,about 23.9±0.2° and about 26.1±0.2°.

In certain embodiments, Compound 1 Pattern 1, optionally as a dihydrate,is characterized by an XRPD pattern comprising the 2theta valuesselected from about 9.6±0.2°, about 12.2±0.2°, about 19.8±0.2°, about21.2±0.2°, about 23.9±0.2° and about 26.1±0.2°.

In certain embodiments, Compound 1 Pattern 1, optionally as a dihydrate,is characterized by an XRPD pattern comprising all or at least 2, 3, 4,5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20° 2thetavalues within±0.4° 2theta of the peaks selected from:

-   -   a. 9.6, 21.3, 19.8, 12.2, 24.0, 26.1, 19.3, 17.6, 28.6, 15.3,        23.2, 20.3, 22.7, 27.1, 21.0, 17.0, 10.1, 16.0, 26.8, 31.4,        13.8, 32.3, 32.9, 20.7, 18.7, and 27.6 °2θ; or    -   b. 9.6, 21.3, 19.8, 12.2, 24.0, 26.1, 19.3, 17.6, 28.6, 15.3,        23.2, 20.3, 22.7, 27.1, 21.0, 17.0, 10.1, 16.0, 26.8, 31.4,        13.8, 32.3, 32.9, 20.7, and 18.7 °2θ; or    -   c. 9.6, 21.3, 19.8, 12.2, 24.0, 26.1, 19.3, 17.6, 28.6, 15.3,        23.2, 20.3, 22.7, 27.1, 21.0, 17.0, 10.1, 16.0, 26.8, 31.4,        13.8, 32.3, 32.9, and 20.7, °2θ; or    -   d. 9.6, 21.3, 19.8, 12.2, 24.0, 26.1, 19.3, 17.6, 28.6, 15.3,        23.2, 20.3, 22.7, 27.1, 21.0, 17.0, 10.1, 16.0, 26.8, 31.4,        13.8, 32.3, and 32.9, °2θ; or    -   e. 9.6, 21.3, 19.8, 12.2, 24.0, 26.1, 19.3, 17.6, 28.6, 15.3,        23.2, 20.3, 22.7, 27.1, 21.0, 17.0, 10.1, 16.0, 26.8, 31.4,        13.8, and 32.3, °2θ; or    -   f. 9.6, 21.3, 19.8, 12.2, 24.0, 26.1, 19.3, 17.6, 28.6, 15.3,        23.2, 20.3, 22.7, 27.1, 21.0, 17.0, 10.1, 16.0, 26.8, 31.4, and        13.8, °2θ; or    -   g. 9.6, 21.3, 19.8, 12.2, 24.0, 26.1, 19.3, 17.6, 28.6, 15.3,        23.2, 20.3, 22.7, 27.1, 21.0, 17.0, 10.1, 16.0, 26.8, and 31.4,        °2θ; or    -   h. 9.6, 21.3, 19.8, 12.2, 24.0, 26.1, 19.3, 17.6, 28.6, 15.3,        23.2, 20.3, 22.7, 27.1, 21.0, 17.0, 10.1, 16.0, and 26.8, °2θ;        or    -   i. 9.6, 21.3, 19.8, 12.2, 24.0, 26.1, 19.3, 17.6, 28.6, 15.3,        23.2, 20.3, 22.7, 27.1, 21.0, 17.0, 10.1, and 16.0, °2θ; or    -   j. 9.6, 21.3, 19.8, 12.2, 24.0, 26.1, 19.3, 17.6, 28.6, 15.3,        23.2, 20.3, 22.7, 27.1, 21.0, 17.0, and 10.1, °2θ; or    -   k. 9.6, 21.3, 19.8, 12.2, 24.0, 26.1, 19.3, 17.6, 28.6, 15.3,        23.2, 20.3, 22.7, 27.1, 21.0, and 17.0, °2θ; or    -   l. 9.6, 21.3, 19.8, 12.2, 24.0, 26.1, 19.3, 17.6, 28.6, 15.3,        23.2, 20.3, 22.7, 27.1, and 21.0, °2θ; or    -   m. 9.6, 21.3, 19.8, 12.2, 24.0, 26.1, 19.3, 17.6, 28.6, 15.3,        23.2, 20.3, 22.7, and 27.1, °2θ; or    -   n. 9.6, 21.3, 19.8, 12.2, 24.0, 26.1, 19.3, 17.6, 28.6, 15.3,        23.2, 20.3, and 22.7, °2θ; or    -   o. 9.6, 21.3, 19.8, 12.2, 24.0, 26.1, 19.3, 17.6, 28.6, 15.3,        23.2, and 20.3, °2θ; or    -   p. 9.6, 21.3, 19.8, 12.2, 24.0, 26.1, 19.3, 17.6, 28.6, 15.3,        and 23.2, °2θ; or    -   q. 9.6, 21.3, 19.8, 12.2, 24.0, 26.1, 19.3, 17.6, 28.6, and        15.3, °2θ; or    -   r. 9.6, 21.3, 19.8, 12.2, 24.0, 26.1, 19.3, 17.6, and 28.6, °2θ;        or    -   s. 9.6, 21.3, 19.8, 12.2, 24.0, 26.1, 19.3, and 17.6, °2θ; or    -   t. 9.6, 21.3, 19.8, 12.2, 24.0, 26.1, and 19.3, °2θ; or    -   u. 9.6, 21.3, 19.8, 12.2, 24.0, and 26.1, °2θ;    -   v. any of the above peak lists wherein the °2θ are±0.1; or    -   w. any of the above peak lists wherein the °2θ are±0.2; or    -   x. any of the above peak lists wherein the °2θ are±0.3.

In one embodiment Compound 1 Pattern 1, optionally as a dihydrate, ischaracterized by an XRPD pattern described above and is furthercharacterized by having no peaks greater than 15% relative intensity inbetween 4 and 9 °2θ. In one embodiment Compound 1 Pattern 1, optionallyas a dihydrate, is characterized by an XRPD pattern described above andis further characterized by having no peaks greater than 10% relativeintensity in between 4 and 9 °2θ. In one embodiment Compound 1 Pattern1, optionally as a dihydrate, is characterized by an XRPD patterndescribed above and is further characterized by having no peaks greaterthan 5% relative intensity in between 4 and 9 °2θ. In one embodimentC0ompound 1 Pattern 1, optionally as a dihydrate, is characterized by anXRPD pattern described above and is further characterized by having nopeaks greater than 3% relative intensity in between 4 and 9 °2θ.

In certain embodiments Compound 1 Pattern 1, optionally as a dihydrate,is used in the preparation of amorphous Compound 1 as a dihydrochloride.For example, the amorphous Compound 1 as a dihydrochloride can be formedby lyophilization or spray drying of Compound 1 Pattern 1, optionally asa dihydrate. This amorphous dihydrochloride may possess higher puritythan if it were directly prepared by known synthetic methods. Inaddition, an advantage of first preparing Compound 1 Pattern 1,optionally as a dihydrate, and then converting it to an amorphicmaterial may be that the total shelf life of the product is increased asa result.

In certain embodiments Compound 1 Pattern 1, optionally as a dihydrate,is used in the preparation of amorphous Compound 1 as a mono-HCl salt.For example, the amorphous Compound 1 as a mono-HCl salt can be formedby dissolving Compound 1 Pattern 1 and modifying the pH of the solutionfollowed by removal of solvent. This amorphous mono-HCl salt may possesshigher purity than if it was directly prepared by known syntheticmethods. In addition, an advantage of first preparing Compound 1 Pattern1, optionally as a dihydrate, and then converting it to an amorphicmaterial may be that the total shelf life of the product is increased.

In certain embodiments Compound 1 Pattern 1, optionally as a dihydrate,is used in the preparation of amorphous Compound 1 as a free base. Thisamorphous free base may possess higher purity than if it was directlyprepared by known synthetic methods. In addition, an advantage of firstpreparing Compound 1 Pattern 1, optionally as a dihydrate, and thenconverting it to an amorphic material may be that the total shelf lifeof the product is increased as a result.

In certain embodiments Compound 1 Pattern 1, optionally as a dihydrate,is used in the preparation of a liquid solution suitable for intravenousadministration of Compound 1. This liquid solution of Compound 1 maypossess higher purity than if it was directly prepared by knownsynthetic methods. In addition, an advantage of first preparing Compound1 Pattern 1, optionally as a dihydrate, and then converting it to aliquid solution may be that the total shelf life of the product isincreased as a result.

In certain embodiments Compound 1 Pattern 1, optionally as a dihydrate,is lyophilized. In these embodiments Compound 1 Pattern 1, optionally asa dihydrate, can optionally be mixed with one or more appropriateexcipients before or after lyophilization. For example, in certainembodiments the invention provides a solid lyophilized compositioncomprising Compound 1 Pattern 1, optionally as a dihydrate. In certainembodiments the invention provides solid lyophilized compositioncomprising Compound 1 Pattern 1, optionally as a dihydrate, mannitol,and citric acid. In certain embodiments the invention provides solidlyophilized composition comprising Compound 1 Pattern 1 dihydrochloride,dihydrate, mannitol, and citric acid.

Non-Limiting Embodiments of the Present Invention

-   -   1. In certain embodiments a crystalline compound is provided of        structure:

which is a dihydrochloride, dihydrate.

Other non-limiting embodiments include:

-   -   2. The crystalline compound of embodiment 1 characterized by an        X-ray powder diffraction (XRPD) pattern comprising at least        three 2theta values selected from 9.6±0.2°, 21.3±0.2°,        19.8±0.2°, 12.2±0.2°, 24.0±0.2°, 26.1±0.2°, 19.3±0.2°,        17.6±0.2°, and 28.6±0.2°.    -   3. The crystalline compound of embodiment 2, wherein the XRPD        pattern comprises at least four 2theta values selected from        9.6±0.2°, 21.3±0.2°, 19.8±0.2°, 12.2±0.2°, 24.0±0.2°, 26.1±0.2°,        19.3±0.2°, 17.6±0.2°, and 28.6±0.2°.    -   4. The crystalline compound of embodiment 2, wherein the XRPD        pattern comprises at least five 2theta values selected from        9.6±0.2°, 21.3±0.2°, 19.8±0.2°, 12.2±0.2°, 24.0±0.2°, 26.1±0.2°,        19.3±0.2°, 17.6±0.2°, and 28.6±0.2°.    -   5. The crystalline compound of embodiment 2, wherein the XRPD        pattern comprises at least six 2theta values selected from        9.6±0.2°, 21.3±0.2°, 19.8±0.2°, 12.2±0.2°, 24.0±0.2°, 26.1±0.2°,        19.3±0.2°, 17.6±0.2°, and 28.6±0.2°.    -   6. The crystalline compound of embodiment 2, wherein the XRPD        pattern comprises at least the 2theta value of 9.6±0.2°.    -   7. The crystalline compound of embodiment 2, wherein the XRPD        pattern comprises at least the 2theta values of 9.6±0.2°,        19.8±0.2°, and 21.3±0.2°.    -   8. In certain embodiments a lyophilized powder prepared from the        crystalline compound of embodiment 1 is provided.    -   9. The lyophilized powder of embodiment 8, wherein the        crystalline compound of embodiment 1 is characterized by an        X-ray powder diffraction (XRPD) pattern comprising at least        three 2theta values selected from 9.6±0.2°, 21.3±0.2°,        19.8±0.2°, 12.2±0.2°, 24.0±0.2°, 26.1±0.2°, 19.3±0.2°,        17.6±0.2°, and 28.6±0.2°.    -   10. The lyophilized powder of embodiment 9, wherein the        crystalline compound of embodiment 1 is characterized by an XRPD        pattern comprising at least four 2theta values selected from        9.6±0.2°, 21.3±0.2°, 19.8±0.2°, 12.2±0.2°, 24.0±0.2°, 26.1±0.2°,        19.3±0.2°, 17.6±0.2°, and 28.6±0.2°.    -   11. The lyophilized powder of embodiment 9, wherein the        crystalline compound of embodiment 1 is characterized by an XRPD        pattern comprising at least five 2theta values selected from        9.6±0.2°, 21.3±0.2°, 19.8±0.2°, 12.2±0.2°, 24.0±0.2°, 26.1±0.2°,        19.3±0.2°, 17.6±0.2°, and 28.6±0.2°.    -   12. The lyophilized powder of embodiment 9, wherein the        crystalline compound of embodiment 1 is characterized by an XRPD        pattern comprising at least six 2theta values selected from        9.6±0.2°, 21.3±0.2°, 19.8±0.2°, 12.2±0.2°, 24.0±0.2°, 26.1±0.2°,        19.3±0.2°, 17.6±0.2°, and 28.6±0.2°.    -   13. The lyophilized powder of embodiment 9 wherein the        crystalline compound of embodiment 1 is characterized by an XRPD        pattern comprising at least the 2theta value of 9.6±0.2°.    -   14. The lyophilized powder of embodiment 9, wherein the        crystalline compound of embodiment 1 is characterized by an XRPD        pattern comprising at least the 2theta values of 9.6±0.2°,        19.8±0.2°, and 21.3±0.2°.    -   15. In certain embodiments a pharmaceutical composition        comprising the crystalline compound of embodiment 1 and a        pharmaceutically acceptable excipient is provided.    -   16. The pharmaceutical composition of embodiment 15, wherein the        pharmaceutical composition is suitable for intravenous delivery.    -   17. The pharmaceutical composition of embodiment 15 comprising        about 200 milligrams to about 600 milligrams of the crystalline        compound of embodiment 1.    -   18. The pharmaceutical composition of embodiment 15 comprising        about 300 milligrams of the crystalline compound of embodiment        1.    -   19. The pharmaceutical composition of embodiment 15 comprising a        dose of about 150 mg/m² to about 350 mg/m² of the crystalline        compound of embodiment 1.    -   20. The pharmaceutical composition of embodiment 15 further        comprising about 300 mg of mannitol and about 76 mg of citric        acid.    -   21. The pharmaceutical composition of embodiment 15 comprising a        dose of about 240 mg/m² of the crystalline compound of        embodiment 1.    -   22. In certain embodiments a pharmaceutically acceptable        reconstituted solution of the lyophilized powder of embodiment 8        is provided.    -   23. The pharmaceutical solution of embodiment 23, wherein the        solution is aqueous.    -   24. The pharmaceutical solution of embodiment 23, wherein the        pharmaceutical composition is suitable for intravenous delivery.    -   25. The pharmaceutical solution of embodiment 23 comprising        about 200 milligrams to about 600 milligrams of the lyophilized        powder of embodiment 8.    -   26. The pharmaceutical solution of embodiment 23 comprising        about 300 milligrams of the lyophilized powder of embodiment 8.    -   27. The pharmaceutical solution of embodiment 23 comprising a        dose of about 150 mg/m² to about 350 mg/m² of the lyophilized        powder of embodiment 8.    -   28. The pharmaceutical solution of embodiment 23 further        comprising about 300 mg of mannitol and about 76 mg of citric        acid.    -   29. The pharmaceutical solution of embodiment 23 further        comprising sodium hydroxide or hydrochloric acid.    -   30. The pharmaceutical solution of embodiment 23 comprising a        dose of about 240 mg/m² of the lyophilized powder of embodiment        8.    -   31. A lyophilized powder comprising a compound of structure:

which is a dihydrochloride, which optionally includes hydrate.

-   -   32. The lyophilized powder of embodiment 31, wherein the        compound is characterized by an X-ray powder diffraction (XRPD)        pattern comprising at least three 2theta values selected from        9.6±0.2°, 21.3±0.2°, 19.8±0.2°, 12.2±0.2°, 24.0±0.2°, 26.1±0.2°,        19.3±0.2°, 17.6±0.2°, and 28.6±0.2°.    -   33. The lyophilized powder of embodiment 31, wherein the        compound is characterized by an XRPD pattern comprising at least        four 2theta values selected from 9.6±0.2°, 21.3±0.2°, 19.8±0.2°,        12.2±0.2°, 24.0±0.2°, 26.1±0.2°, 19.3±0.2°, 17.6±0.2°, and        28.6±0.2°.    -   34. The lyophilized powder of embodiment 31, wherein the        compound is characterized by an XRPD pattern comprising at least        five 2theta values selected from 9.6±0.2°, 21.3±0.2°, 19.8±0.2°,        12.2±0.2°, 24.0±0.2°, 26.1±0.2°, 19.3±0.2°, 17.6±0.2°, and        28.6±0.2°.    -   35. The lyophilized powder of embodiment 31, wherein the        compound is characterized by an XRPD pattern comprising at least        six 2theta values selected from 9.6±0.2°, 21.3±0.2°, 19.8±0.2°,        12.2±0.2°, 24.0±0.2°, 26.1±0.2°, 19.3±0.2°, 17.6±0.2°, and        28.6±0.2°.    -   36. The lyophilized powder of embodiment 31, wherein the        compound is characterized by an XRPD pattern comprising at least        the 2theta value of 9.6±0.2°.    -   37. The lyophilized powder of embodiment 31, wherein the        compound is characterized by an XRPD pattern comprising at least        the 2theta values of 9.6±0.2°, 19.8±0.2°, and 21.3±0.2°.

Formulation of Compound 1 Pattern 1

In certain embodiments Compound 1 Pattern 1 in the form of adihydrochloride, dihydrate in a formulation is provided. In certainembodiments the formulation comprises about 300-400 mg, about 350-400mg, about 360-380 mg, or more particularly about 373 mg, of Compound 1Pattern 1 dihydrochloride, dihydrate, about 50-100, about 60-80, or moreparticularly about 76 mg of citric acid monohydrate, and about 250-350mg, about 280-320 mg, or more particularly about 300 mg of mannitol, andoptionally sodium hydroxide and/or hydrochloric acid to adjust the pH asdesired. In particular, 373 mg of the dihydrochloride, dihydrate willprovide 300 mg of Trilaiclib free base.

In certain embodiments the formulation is lyophilized after Compound 1Pattern 1 in the form of a dihydrochloride, dihydrate is mixed withcitric acid and mannitol. In certain embodiments the formulationcomprises lyophilized Compound 1 Pattern 1 in the form of adihydrochloride, dihydrate mixed with citric acid and mannitol, whereinthe citric acid and mannitol are added after Compound 1 Pattern 1 in theform of a dihydrochloride, dihydrate is lyophilized.

In certain embodiments Compound 1 Pattern 1 in the form of adihydrochloride in a formulation is provided. In certain embodiments theformulation comprises about 300-400 mg, about 350-400 mg, or about360-380 mg, or more particularly about 373 mg, of Compound 1 Pattern 1dihydrochloride, dihydrate, about 50-100, about 60-80, or moreparticularly about 76 mg of citric acid monohydrate, and about 250-350mg, about 280-320 mg or more particularly about 300 mg of mannitol, andoptionally sodium hydroxide and/or hydrochloric acid to adjust the pH asdesired for reconstitution.

In certain embodiments this formulation is sterile.

In certain embodiments the formulation is lyophilized after Compound 1Pattern 1 in the form of a dihydrochloride, dihydrate is mixed withcitric acid and mannitol. In certain embodiments the formulationcomprises lyophilized Compound 1 Pattern 1 in the form of adihydrochloride mixed with citric acid and mannitol, wherein the citricacid and mannitol are added after Compound 1 Pattern 1 in the form of adihydrochloride, dihydrate is lyophilized.

In certain embodiments a formulation is provided wherein the formulationis prepared by lyophilizing about 300-400 mg, about 350-400 mg, or about360-380 mg, or more particularly about 373 mg, of Compound 1 Pattern 1dihydrochloride, dihydrate, about 50-100, about 60-80, or moreparticularly about 76 mg citric acid monohydrate, and about 250-350 mg,about 280-320 mg or more particularly about 300 mg of mannitol, andoptionally sodium hydroxide and/or hydrochloric acid to adjust the pH asdesired. In another embodiment the formulation results from lyophilizingCompound 1 Pattern 1 after it is mixed with about 50-100 mg, about 60-80mg, or more particularly about 76 mg of citric acid monohydrate, andabout 280-320 mg, or more particularly about 300 mg of mannitol.

Any of the formulations described above can be reconstituted with about5-100, 5-50, 10-30, about 10-25, or more particularly about 19.5 mL ofsterile water, phosphate buffered saline, dilute sugar, or anothersaline solution. In one non-limiting embodiment the reconstitutionsolution is a sterile sodium chloride solution, for example, comprisingabout 0.9% NaCl or a sterile sugar solution, for example, of about 5%dextrose solution. The resulting reconstituted solution will have anyamount of trilaciclib needed for the intended purpose, such as forexample, between about 5-50 mg/mL, about 10-25 mg/ml or even about 15mg/mL of trilaciclib. In certain embodiments the resulting solution isthen diluted prior to parenteral administration, such as intravenousdelivery.

Non-limiting examples of typical quality standards and pharmaceuticalfunction are provided in the table below.

Quantity (mg) per Pharmaceutical Quality Components Vial^(a) FunctionStandards trilaciclib^(b) 300 Active Internal Mannitol 300 Bulking agentUSP/Ph. Eur. Citric acid 75.6 Buffer USP/Ph. Eur. monohydrate Sodiumhydroxide q.s. pH adjustment NF/Ph. Eur. Hydrochloric acid q.s. pHadjustment^(c) NF/Ph. Eur. Water for Injection^(d) q.s. Solvent USP/Ph.Eur. Nitrogen^(e) N/A Process aide NF NF = National Formulary; USP =United States Pharmacopoeia; Ph. Eur. = European Pharmacopoeia; N/A =not applicable; q.s. = quantity sufficient ^(a)The target weight ofsolution filled into vials prior to lyophilization is 12.252 g (12 mL).^(b)Amount in terms of free base (equivalent to 373 mg of Compound 1Pattern 1 dihydrochloride, dihydrate). ^(c)May be used if necessary, toadjust the bulk solution pH. ^(d)Essentially removed duringlyophilization. ^(e)Nitrogen is used as an inert gas at the end of thelyophilization process for vacuum adjustment.

The resulting formulation can be supplied in any appropriate container,for example in a 20 mL (Type 1) clear glass vial, sealed with a 20 mmgray chlorobutyl rubber stopper and secured with a 20 mm aluminumoverseal with a plastic flip-off seal.

In certain embodiments the solid formulation is reconstituted with about5-100, 5-50, 10-30, about 10-25, or more particularly about 19.5 mL ofabout 0.9% NaCl or a sterile sugar solution, for example, of about 5%dextrose solution, sodium hydroxide and/or hydrochloric acid is added toadjust pH, and then the reconstituted solution is diluted to theappropriate dose for administration to a human in need thereof, forexample at a dosage of about 200-300 mg/m², for example 240 mg/m².

Chemical Description and Terminology

Compounds are described using standard nomenclature. Unless definedotherwise, all technical and scientific terms used herein have the samemeaning as is commonly understood by one of skill in the art to whichthis invention belongs.

The terms “a” and “an” do not denote a limitation of quantity, butrather denote the presence of at least one of the referenced item. Theterm “or” means “and/or”. Recitation of ranges of values are merelyintended to serve as a shorthand method of referring individually toeach separate value falling within the range, unless otherwise indicatedherein, and each separate value is incorporated into the specificationas if it were individually recited herein. The endpoints of all rangesare included within the range and independently combinable.

All methods described herein can be performed in any suitable orderunless otherwise indicated herein or otherwise clearly contradicted bycontext. The use of any and all examples, or exemplary language (e.g.,“such as”), is intended merely for illustration and does not pose alimitation on the scope of the invention unless otherwise claimed.

An “active agent” is a compound (including a compound disclosed herein),element, or mixture that when administered to a patient, alone or incombination with another compound, element, or mixture, confers,directly or indirectly, a physiological effect on the patient. Theindirect physiological effect may occur via a metabolite or otherindirect mechanism.

“Deuteration” and “deuterated” means that a hydrogen is replaced by adeuterium such that the deuterium exists over natural abundance and isthus “enriched”. An enrichment of 50% means that rather than hydrogen atthe specified position the deuterium content is 50%. For clarity, it isconfirmed that the term “enriched” as used herein does not meanpercentage enriched over natural abundance. In other embodiments, therewill be at least 80%, at least 90%, or at least 95% deuterium enrichmentat the specified deuterated position or positions. In other embodimentsthere will be at least 96%, at least 97%, at least 98%, or at least 99%deuterium enrichment at the specified deuterated position or positionsindicated. In the absence of indication to the contrary, the enrichmentof deuterium in the specified position of the compound described hereinis at least 90%.

A “dosage form” means a unit of administration of an active agent.Non-limiting examples of dosage forms include tablets, capsules,injections, suspensions, liquids, intravenous fluids, emulsions, creams,ointments, suppositories, inhalable forms, transdermal forms, and thelike.

“Pharmaceutical compositions” are compositions comprising at least oneactive agent, such as a compound or salt of one of the active compoundsdisclosed herein, and at least one other substance, such as a carrier.Pharmaceutical compositions optionally contain more than one activeagent. “Pharmaceutical combinations” or “combination therapy” refers tothe administration of at least two active agents, and in one embodiment,three or four or more active agents which may be combined in a singledosage form or provided together in separate dosage forms optionallywith instructions that the active agents are to be used together totreat a disorder.

“Pharmaceutically acceptable salts” includes derivatives of thedisclosed compounds in which the parent compound is modified by makinginorganic and organic, suitably non-toxic, acid or base addition saltsthereof. The salts of the present compounds can be synthesized from aparent compound that contains a basic or acidic moiety by conventionalchemical methods. Generally, such salts can be prepared by reacting freeacid forms of these compounds with a stoichiometric amount of theappropriate base (such as Na, Ca, Mg, or K hydroxide, carbonate,bicarbonate, or the like), or by reacting free base forms of thesecompounds with a stoichiometric amount of the appropriate acid. Suchreactions are typically carried out in water or in an organic solvent,or in a mixture of the two. The pharmaceutically acceptable salt can bein the form of a pure crystal, or single morphic form, or can be used innon-crystalline or amorphic, glassy, or vitreous form, or a mixturethereof. In an alternative embodiment, the active compound can beprovided in the form of a solvate.

Examples of pharmaceutically acceptable salts include, but are notlimited to, mineral or organic acid salts of basic residues such asamines; alkali or organic salts of acidic residues such as carboxylicacids; and the like. The pharmaceutically acceptable salts include theconventional non-toxic salts and the quaternary ammonium salts of theparent compound formed, for example, from non-toxic inorganic or organicacids. For example, conventional non-toxic acid salts include thosederived from inorganic acids such as hydrochloric, hydrobromic,sulfuric, sulfamic, phosphoric, nitric and the like; and the saltsprepared from organic acids such as acetic, propionic, succinic,glycolic, stearic, lactic, malic, tartaric, citric, ascorbic, pamoic,maleic, hydroxymaleic, phenylacetic, glutamic, benzoic, salicylic,mesylic, esylic, besylic, sulfanilic, 2-acetoxybenzoic, fumaric,toluenesulfonic, methanesulfonic, ethane disulfonic, oxalic, isethionic,HOOC—(CH₂)n-COOH where n is 0-4, and the like. Lists of additionalsuitable salts may be found, e.g., in Remington's PharmaceuticalSciences, 17th ed., Mack Publishing Company, Easton, Pa., p. 1418(1985).

The term “carrier” means a diluent, excipient, or vehicle with which anactive compound is provided.

A “pharmaceutically acceptable excipient” means an excipient that isuseful in preparing a pharmaceutical composition/combination that isgenerally safe, is sufficiently non-toxic, and neither biologically norotherwise undesirable. A “pharmaceutically acceptable excipient” as usedin the present application includes both one and more than one suchexcipient.

A “patient” or “host” is a human or non-human animal, including, but notlimited to, simian, avian, feline, canine, bovine, equine or porcine inneed of medical treatment. Medical treatment can include treatment of anexisting condition, such as a disease or disorder, or a prophylactic ordiagnostic treatment. In a particular embodiment, the patient or host isa human patient. The patient such as a host can be treated for anycondition that is responsive to trilaciclib, including formyelopreservation or as an antineoplastic agent.

The term “isolated” as used herein refers to the material insubstantially pure form. An isolated compound does not have anothercomponent that materially affects the properties of the compound. Inparticular embodiments, an isolated form is at least 50, 60, 70, 80, 90,95, 98 or 99% pure.

Pharmaceutical Compositions and Dosage Forms

In one aspect of the invention a formulation is provided wherein theformulation is prepared by lyophilizing about 300 to 400 mg, about350-400 mg or more particularly about 373 mg of Compound 1 Pattern 1dihydrochloride, dihydrate (equivalent to about 300 mg of trilaciclibfree base) and mixing it with about 50-100 mg, about 60-80 mg, or moreparticularly about 76 mg of citric acid monohydrate, and about 250-350mg, about 280-320 mg or more particularly about 300 mg of mannitol, andoptionally sodium hydroxide and/or hydrochloric acid to adjust the pH asdesired. In another embodiment the formulation results from lyophilizingCompound 1 Pattern 1 after it is mixed with about 50-100 mg, about 60-80mg, or more particularly about 76 mg of citric acid monohydrate, andabout 250-350 mg, about 280-320 mg or more particularly about 300 mg ofmannitol, and optionally sodium hydroxide and/or hydrochloric acid toadjust the pH as desired.

This formulation can be reconstituted with about 10-30, or moreparticularly about 5-100, 5-50, 10-30, about 10-25, or more particularlyabout 19.5 mL of sterile water, phosphate buffered saline, dilute sugar,or another saline solution. In one non-limiting embodiment thereconstitution solution is a sterile sodium chloride solution, forexample, comprising about 0.9% NaCl or a sterile sugar solution, forexample, of about 5% dextrose solution. The resulting reconstitutedsolution will have any amount of trilaciclib needed for the intendedpurpose, such as for example, between about 5-50 mg/mL, about 10-25mg/ml or even about 15 mg/mL of trilaciclib. In certain embodiments theresulting solution is then diluted prior to parenteral administration,such as intravenous delivery.

The isolated Compound 1 Pattern 1 described herein, or an alternativesalt, isotopic analog, or prodrug can be administered in an effectiveamount to a host to treat any of the disorders described herein usingany suitable approach which achieves the desired therapeutic result. Theamount and timing of the isolated Compound 1 Pattern 1 administeredwill, of course, be dependent on the host being treated, theinstructions of the supervising medical specialist, on the time courseof the exposure, on the manner of administration, on the pharmacokineticproperties of the particular active compound, and on the judgment of theprescribing physician. Thus, because of host to host variability, thedosages given below are a guideline and the physician can titrate dosesof the compound to achieve the treatment that the physician considersappropriate for the host. In considering the degree of treatmentdesired, the physician can balance a variety of factors such as age andweight of the host, presence of preexisting disease, as well as presenceof other diseases.

The pharmaceutical composition may be formulated as any pharmaceuticallyuseful form, and typically a parenteral formulation, such as anintravenous, intramuscular, subcutaneous, or intradermal formulation.Other alternative formulations include oral, transdermal, or intranasalformulations. For example the pharmaceutical composition may be in theform of an I.V. bag a vial for injection, a pill, a capsule, a tablet, atransdermal patch, a subcutaneous patch, a dry powder, an inhalationformulation, in a medical device, suppository, buccal, or sublingualformulation. Some dosage forms, such as tablets and capsules, aresubdivided into suitably sized unit doses containing appropriatequantities of the active components, e.g., an effective amount toachieve the desired purpose.

In certain embodiments the pharmaceutical composition is in a dosageform that contains from about 50 mg/m² to about 800 mg/m², from about100 mg/m² to about 600 mg/m², from about 100 mg/m² to about 500 mg/m²,from about 100 mg/m² to about 400 mg/m², from about 100 mg/m² to about350 mg/m², from about 150 mg/m² to about 350 mg/m², from about 200 mg/m²to about 350 mg/m², or from about 200 mg/m² to about 300 mg/m². In oneembodiment the pharmaceutical composition is in a dosage form thatcontains about 240 mg/m².

The therapeutically effective dosage of the isolated Compound 1 Pattern1 described herein will be determined by the health care practitionerdepending on the condition, size and age of the patient as well as theroute of delivery. In one non-limited embodiment, a dosage from about0.1 to about 200 mg/kg has therapeutic efficacy, with all weights beingcalculated based upon the weight of the active compound. In someembodiments, the dosage may be the amount of the isolated Compound 1Pattern 1 needed to provide a serum concentration of the active compoundof up to about 10 nM, 50 nM, 100 nM, 200 nM, 300 nM, 400 nM, 500 nM, 600nM, 700 nM, 800 nM, 900 nM, 1 μM, 5 μM, 10 μM, 20 μM, 30 μM, or 40 μM.

In certain embodiments the pharmaceutical composition is in a dosageform that contains from about 0.1 mg to about 2000 mg, from about 10 mgto about 1000 mg, from about 100 mg to about 800 mg, or from about 200mg to about 600 mg of the active compound and optionally from about 0.1mg to about 2000 mg, from about 10 mg to about 1000 mg, from about 100mg to about 800 mg, or from about 200 mg to about 600 mg, for examplefrom about 300 mg to about 400 mg of the isolated Compound 1 Pattern 1,measured alternatively either as the free base or its salt, in a unitdosage form, for example for parenteral deliver such as an I.V. bag.Examples are dosage forms with at least 5, 10, 15, 20, 25, 50, 100, 200,250, 300, 400, 500, 600, 700, or 750 mg of active compound, or its salt.The pharmaceutical composition may also include a molar ratio of theisolated Compound 1 Pattern 1 and an additional active agent, in a ratiothat achieves the desired results.

The isolated Compound 1 Pattern 1 disclosed herein or used as describedherein may be administered parenterally, intravaneously, orally,topically, by inhalation or spray, sublingually, via implant, includingocular implant, transdermally, via buccal administration, rectally,intramuscular, inhalation, intra-aortal, intracranial, subdermal,intraperitioneal, subcutaneous, transnasal, sublingual, or rectal or byother means, in dosage unit formulations containing conventionalpharmaceutically acceptable carriers.

In accordance with the presently disclosed methods, a parenteral dosageform can be made from Compound 1 Pattern 1, for example Compound 1Pattern 1 dihydrochloride, dihydrate. This dosage form can include anypharmaceutically acceptable excipient, for example a liquid excipient.Non-limiting examples of liquid excipients include phosphate bufferedsaline, unbuffered or buffered saline (e.g. NaCl solution withoutbuffering agents), a sugar solution (e.g. a solution of dextrose, or acombination thereof as desired. When being prepared for parenteraldosing Compound 1 Pattern 1, for example Compound 1 Pattern 1dihydrochloride, dihydrate, may be dissolved in a concentrated solutionof an appropriate liquid excipient. This concentrated solution ofCompound 1 may then be diluted with the same or different liquidexcipient to the appropriate dose for treating a human in need thereof.In certain embodiments the pH of the solution (either before or afterdilution) is adjusted with a pH adjusting reagent (for example HCl orNaOH). In certain embodiments an additional therapeutic ornon-therapeutic agent is added to the solution before administration orto improve shelf life.

In accordance with the presently disclosed methods, an oraladministration can be in any desired form in which the isolated Compound1 Pattern 1 is stable as a solid. In certain embodiments, the isolatedCompound 1 Pattern 1 is delivered in a solid microparticle ornanoparticle. When administered through inhalation the isolated Compound1 Pattern 1 may be in the form of a plurality of solid particles ordroplets having any desired particle size, and for example, from about0.01, 0.1 or 0.5 to about 5, 10, 20 or more microns, and optionally fromabout 1 to about 2 microns. The isolated Compound 1 Pattern 1 asdisclosed in the present invention has good pharmacokinetic andpharmacodynamics properties, for instance when administered by the oralor intravenous routes.

The pharmaceutical formulations can comprise the isolated Compound 1Pattern 1 described herein or an alternative pharmaceutically acceptablesalt thereof, in any pharmaceutically acceptable carrier.

Carriers include excipients and diluents and must be of sufficientlyhigh purity and sufficiently low toxicity to render them suitable foradministration to the patient being treated. The carrier can be inert orit can possess pharmaceutical benefits of its own. The amount of carrieremployed in conjunction with the compound is sufficient to provide apractical quantity of material for administration per unit dose of thecompound.

Classes of carriers include, but are not limited to binders, bufferingagents, coloring agents, diluents, di sintegrants, emulsifiers,flavorants, glidents, lubricants, preservatives, stabilizers,surfactants, tableting agents, and wetting agents. Some carriers may belisted in more than one class, for example vegetable oil may be used asa lubricant in some formulations and a diluent in others. Exemplarypharmaceutically acceptable carriers include sugars, starches,celluloses, powdered tragacanth, malt, gelatin; talc, and vegetableoils. Optional active agents may be included in a pharmaceuticalcomposition, which do not substantially interfere with the activity ofthe compound of the present invention.

Depending on the intended mode of administration, the pharmaceuticalcompositions can be in the form of solid form or a semi-solid dosageform that the isolated Compound 1 Pattern 1 is stable in, such as, forexample, tablets, suppositories, pills, capsules, powders, or the like,preferably in unit dosage form suitable for single administration of aprecise dosage. The compositions will include an effective amount of theselected drug in combination with a pharmaceutically acceptable carrierand, in addition, can include other pharmaceutical agents, adjuvants,diluents, buffers, and the like.

Thus, the compositions of the disclosure can be administered aspharmaceutical formulations including those suitable for oral (includingbuccal and sub-lingual), rectal, nasal, topical, pulmonary, vaginaladministration or in a form suitable for administration by inhalation orinsufflation. The preferred manner of administration is oral using aconvenient daily dosage regimen which can be adjusted according to thedegree of affliction. For solid compositions, conventional nontoxicsolid carriers include, for example, pharmaceutical grades of mannitol,lactose, starch, magnesium stearate, sodium saccharin, talc, cellulose,glucose, sucrose, magnesium carbonate, and the like.

In yet another embodiment is the use of permeation enhancer excipientsincluding polymers such as: polycations (chitosan and its quaternaryammonium derivatives, poly-L-arginine, aminated gelatin); polyanions(N-carboxymethyl chitosan, poly-acrylic acid); and, thiolated polymers(carboxymethyl cellulose-cysteine, polycarbophil-cysteine,chitosan-thiobutylamidine, chitosan-thioglycolic acid,chitosan-glutathione conjugates).

For oral administration, the composition will generally take the form ofa tablet or capsule. Tablets and capsules are preferred oraladministration forms. Tablets and capsules for oral use can include oneor more commonly used carriers such as lactose and corn starch.Lubricating agents, such as magnesium stearate, are also typicallyadded. Typically, the compositions of the disclosure can be combinedwith an oral, non-toxic, pharmaceutically acceptable, inert carrier suchas lactose, starch, sucrose, glucose, methyl cellulose, magnesiumstearate, dicalcium phosphate, calcium sulfate, mannitol, sorbitol andthe like. Moreover, when desired or necessary, suitable binders,lubricants, disintegrating agents, and coloring agents can also beincorporated into the mixture. Suitable binders include starch, gelatin,natural sugars such as glucose or beta-lactose, corn sweeteners, naturaland synthetic gums such as acacia, tragacanth, or sodium alginate,carboxymethylcellulose, polyethylene glycol, waxes, and the like.Lubricants used in these dosage forms include sodium oleate, sodiumstearate, magnesium stearate, sodium benzoate, sodium acetate, sodiumchloride, and the like. Disintegrators include, without limitation,starch, methyl cellulose, agar, bentonite, xanthan gum, and the like.

In addition to the active compounds or their salts, the pharmaceuticalformulations can contain other additives, such as pH-adjustingadditives. In particular, useful pH-adjusting agents include acids, suchas hydrochloric acid, bases or buffers, such as sodium lactate, sodiumacetate, sodium phosphate, sodium citrate, sodium borate, or sodiumgluconate. Further, the formulations can contain antimicrobialpreservatives. Useful antimicrobial preservatives include methylparaben,propylparaben, and benzyl alcohol. An antimicrobial preservative istypically employed when the formulations is placed in a vial designedfor multi-dose use. The pharmaceutical formulations described herein canbe lyophilized using techniques well known in the art.

For oral administration a pharmaceutical composition can take the formof a tablet, pill, capsule, powder, and the like. Tablets containingvarious excipients such as sodium citrate, calcium carbonate and calciumphosphate may be employed along with various disintegrants such asstarch (e.g., potato or tapioca starch) and certain complex silicates,together with binding agents such as polyvinylpyrrolidone, sucrose,gelatin and acacia. Additionally, lubricating agents such as magnesiumstearate, sodium lauryl sulfate, and talc are often very useful fortableting purposes. Solid compositions of a similar type may be employedas fillers in soft and hard-filled gelatin capsules.

Pharmaceutical formulations also are provided which provide a controlledrelease of a compound described herein, including through the use of adegradable polymer, as known in the art.

The term “pharmaceutically acceptable salts” as used herein refers tothose salts which are, within the scope of sound medical judgment,suitable for use in contact with hosts (e.g., human hosts) without unduetoxicity, irritation, allergic response, and the like, commensurate witha reasonable benefit/risk ratio, and effective for their intended use,as well as the zwitterionic forms, where possible, of the compounds ofthe presently disclosed host matter.

In an alternative embodiment Compound 1 Pattern 1 is not a HCl salt, butis instead a salt described below.

In one embodiment the additional therapeutic agent described in theCombination Section below is administered as a pharmaceuticallyacceptable salt, for example, a salt described below.

Thus, the term “salts” refers to the relatively non-toxic, inorganic andorganic acid addition salts of the presently disclosed compounds. Thesesalts can be prepared during the final isolation and purification of thecompounds or by separately reacting the purified compound in its freebase form with a suitable organic or inorganic acid and isolating thesalt thus formed. Basic compounds are capable of forming a wide varietyof different salts with various inorganic and organic acids. Acidaddition salts of the basic compounds are prepared by contacting thefree base form with a sufficient amount of the desired acid to producethe salt in the conventional manner. The free base form can beregenerated by contacting the salt form with a base and isolating thefree base in the conventional manner. The free base forms may differfrom their respective salt forms in certain physical properties such assolubility in polar solvents. Pharmaceutically acceptable base additionsalts may be formed with metals or amines, such as alkali and alkalineearth metal hydroxides, or of organic amines. Examples of metals used ascations, include, but are not limited to, sodium, potassium, magnesium,calcium, and the like. Examples of suitable amines include, but are notlimited to, N,N′-dibenzylethylenediamine, chloroprocaine, choline,diethanolamine, ethylenediamine, N-methylglucamine, and procaine. Thebase addition salts of acidic compounds are prepared by contacting thefree acid form with a sufficient amount of the desired base to producethe salt in the conventional manner. The free acid form can beregenerated by contacting the salt form with an acid and isolating thefree acid in a conventional manner. The free acid forms may differ fromtheir respective salt forms somewhat in certain physical properties suchas solubility in polar solvents.

Salts can be prepared from inorganic acids sulfate, pyrosulfate,bisulfate, sulfite, bisulfite, nitrate, phosphate,monohydrogenphosphate, dihydrogenphosphate, metaphosphate,pyrophosphate, chloride, bromide, iodide such as hydrochloric, nitric,phosphoric, sulfuric, hydrobromic, hydriodic, phosphorus, and the like.Representative salts include the hydrobromide, hydrochloride, sulfate,bisulfate, nitrate, acetate, oxalate, valerate, oleate, palmitate,stearate, laurate, borate, benzoate, lactate, phosphate, tosylate,citrate, maleate, fumarate, succinate, tartrate, naphthylate mesylate,glucoheptonate, lactobionate, laurylsulphonate and isethionate salts,and the like. Salts can also be prepared from organic acids, such asaliphatic mono- and dicarboxylic acids, phenyl-substituted alkanoicacids, hydroxy alkanoic acids, alkanedioic acids, aromatic acids,aliphatic and aromatic sulfonic acids, etc. and the like. Representativesalts include acetate, propionate, caprylate, isobutyrate, oxalate,malonate, succinate, suberate, sebacate, fumarate, maleate, mandelate,benzoate, chlorobenzoate, methylbenzoate, dinitrobenzoate, phthalate,benzenesulfonate, toluenesulfonate, phenylacetate, citrate, lactate,maleate, tartrate, methanesulfonate, and the like.

Pharmaceutically acceptable salts can include cations based on thealkali and alkaline earth metals, such as sodium, lithium, potassium,calcium, magnesium and the like, as well as non-toxic ammonium,quaternary ammonium, and amine cations including, but not limited to,ammonium, tetramethylammonium, tetraethylammonium, methylamine,dimethylamine, trimethylamine, triethylamine, ethylamine, and the like.Also contemplated are the salts of amino acids such as arginate,gluconate, galacturonate, and the like. See, for example, Berge et al.,J. Pharm. Sci., 1977, 66, 1-19, which is incorporated herein byreference.

Formulations suitable for rectal administration are typically presentedas unit dose suppositories. These may be prepared by admixing the activedisclosed compound with one or more conventional solid carriers, forexample, cocoa butter, and then shaping the resulting mixture.

Formulations suitable for topical application to the skin preferablytake the form of an ointment, cream, lotion, paste, gel, spray, aerosol,or oil, which maintain the stability of the isolated Compound 1 Pattern1. Carriers which may be used include petroleum jelly, lanoline,polyethylene glycols, alcohols, transdermal enhancers, and combinationsof two or more thereof.

Formulations suitable for transdermal administration may be presented asdiscrete patches adapted to remain in intimate contact with theepidermis of the recipient for a prolonged period of time. Formulationssuitable for transdermal administration may also be delivered byiontophoresis (see, for example, Pharmaceutical Research 3 (6):318(1986)) and typically take the form of an optionally buffered aqueoussolution of the active compound. In one embodiment, microneedle patchesor devices are provided for delivery of drugs across or into biologicaltissue, particularly the skin. The microneedle patches or devices permitdrug delivery at clinically relevant rates across or into skin or othertissue barriers, with minimal or no damage, pain, or irritation to thetissue.

Formulations suitable for administration to the lungs can be deliveredby a wide range of passive breath driven and active power drivensingle/-multiple dose dry powder inhalers (DPI). The devices mostcommonly used for respiratory delivery include nebulizers, metered-doseinhalers, and dry powder inhalers. Several types of nebulizers areavailable, including jet nebulizers, ultrasonic nebulizers, andvibrating mesh nebulizers. Selection of a suitable lung delivery devicedepends on parameters, such as nature of the drug and its formulation,the site of action, and pathophysiology of the lung.

Advantageous Pharmaceutical Form of Trilaciclib for Effective Therapy

In certain embodiments the morphic form of Compound 1, for exampleCompound 1 dihydrochloride, dihydrate, is used to produce an intravenousformulation as a myelopreservation agent or alternatively when givenalong with chemotherapy to treat cancer optionally along with thestandard of care for the cancer being treated. The standard of care fortreating a particular cancer includes the use of therapies that areapproved by a regulatory agency, for example the U.S. Food and DrugAdministration (FDA), European Medicines Agency (EMA), or the ChineseNational Medical Products Administration (NMPA) for the cancer beingtreated. In other embodiments a pharmaceutical formulation as describedherein is selected and managed by the patient's healthcare practitioner.

(i) Myelopreseryation

When used as a myelopreservation agent, it can be administered, forexample as taught in WO 2014/144326, WO 2016/040848, WO 2018/106729, orPCT/US20/38557. For example, in certain embodiments Compound 1 Pattern 1dihydrochloride, dihydrate or an intravenous solution prepared from itis administered on each day of a treatment cycle on which chemotherapyis administered. For example, if the chemotherapy is given on day 1, day2, and/or day 3 of a treatment cycle then the intravenous solution isalso administered on day 1, day 2, and/or day 3 of the treatment cycle(for example on day 1 or day 1 and day 2 of the treatment cycle). Inanother embodiment the intravenous solution is administered on day 1 andday 8 of a 21 day treatment cycle, and as non-limiting examples, incombination with gemcitabine, carboplatin, or topotecan.

In certain embodiments, the isolated Compound 1 Pattern 1 of the presentinvention decreases the effect of chemotherapeutic agent toxicity onCDK4/6 replication dependent healthy cells, such as hematopoietic stemcells and hematopoietic progenitor cells (together referred to asHSPCs), and/or renal epithelial cells, in subjects, typically humans,that will be, are being, or have been exposed to the chemotherapeuticagent (typically a DNA-damaging agent).

In one embodiment, the subject has been exposed to a chemotherapeuticagent, and, using the intravenous solution described herein, thesubject's CDK4/6-replication dependent healthy cells are placed in G1arrest following exposure in order to mitigate, for example, DNA damage.In one embodiment, the compound is administered at least ½ hour, atleast 1 hour, at least 2 hours, at least 3 hours, at least 4 hours, atleast 5 hours, at least 6 hours, at least 7 hours, at least 8 hours, atleast 10 hours, at least 12 hours, at least 14 hours, at least 16 hours,at least 18 hours, at least 20 hours or more post chemotherapeutic agentexposure. In an alternative embodiment, Compound 1 Pattern 1dihydrochloride, dihydrate or an intravenous solution prepared from itis administered at least ½ hour, at least 1 hour, at least 2 hours, atleast 3 hours, at least 4 hours, at least 5 hours, at least 6 hours, atleast 7 hours, at least 8 hours, at least 10 hours, at least 12 hours,at least 14 hours, at least 16 hours, at least 18 hours, at least 20hours or more prior to chemotherapeutic agent exposure. In certainembodiments Compound 1 Pattern 1 dihydrochloride, dihydrate or anintravenous solution prepared from it is administered about 4 hoursbefore chemotherapy.

In one embodiment, an intravenous solution prepared from Compound 1Pattern 1 dihydrochloride, dihydrate is administered in combination witha chemotherapeutic agent including but not limited to a treatmentregimen wherein the chemotherapeutic agent is administered: on day 1-3every 21 days; on days 1-3 every 28 days; on day 1 every 3 weeks; on day1, day 8, and day 15 every 28 days, on day 1 and day 8 every 28 days; ondays 1 and 8 every 21 days; on days 1-5 every 21 days; 1 day a week for6-8 weeks; on days 1, 22, and 43; days 1 and 2 weekly; days 1-4 and22-25; days 1-4, 22-25, and 43-46; and similar type-regimens, whereinthe CDK4/6-replication dependent cells are arrested at the G1 phaseduring chemotherapeutic agent exposure.

In one embodiment, the isolated Compound 1 Pattern 1 can allow for doseintensification (e.g., more therapy can be given in a fixed period oftime) in medically related chemotherapies, which will translate tobetter efficacy. Therefore, the presently disclosed methods can resultin chemotherapy regimens that are less toxic and more effective.

In some embodiments, the use of the isolated Compound 1 Pattern 1described herein may result in reduced or substantially limitedoff-target effects, for example, related to inhibition of kinases otherthan CDK4 and/or CDK6 such as CDK2. Furthermore, in certain embodiments,the use of the isolated Compound 1 Pattern 1 described herein should notinduce cell cycle arrest in CDK4/6 replication independent cells.

In some embodiments, the use of the isolated Compound 1 Pattern 1described herein reduces the risk of undesirable off-target effectsincluding, but not limited to, long term toxicity, anti-oxidant effects,and estrogenic effects. Anti-oxidant effects can be determined bystandard assays known in the art. For example, a compound with nosignificant anti-oxidant effects is a compound that does notsignificantly scavenge free-radicals, such as oxygen radicals. Theanti-oxidant effects of a compound can be compared to a compound withknown anti-oxidant activity, such as genistein. Thus, a compound with nosignificant anti-oxidant activity can be one that has less than about 2,3, 5, 10, 30, or 100 fold anti-oxidant activity relative to genistein.Estrogenic activities can also be determined via known assays. Forinstance, a non-estrogenic compound is one that does not significantlybind and activate the estrogen receptor. A compound that issubstantially limited estrogenic effects can be one that has less thanabout 2, 3, 5, 10, 20, or 100-fold estrogenic activity relative to acompound with estrogenic activity, e.g., genistein.

Compound 1 Pattern 1, for example Compound 1 Pattern 1 dihydrochloride,dihydrate, or a formulation produced from it may be administeredparenterally, for example, intravenously, to a patient prior toadministration of an immune-response inducing chemotherapy such as anICD-inducing chemotherapy. In some embodiments, Compound 1 isadministered up to about 24 hours or less, or up to about 20, 15, 10, 5,or 4 hours or less for example about 30-60 minutes or less, prior toadministration of the chemotherapy. In some embodiments, Compound 1 isadministered approximately about 22 to 26 hours before administration ofthe chemotherapy, and again about 4 hours or less, for example about30-60 minutes or less, prior to administration of the chemotherapy. Insome embodiments, the dose of Compound 1 administered is between about180 and about 280 mg/m². For example, the dose is up to about 100, 125,150, 180, 185, 190, 195, 200, 205, 210, 215, 220, 225, 230, 235, 240,245, 250, 255, 260, 265, 270, 275, or 280 mg/m² or any dose in betweenthese numbers as determined desirable by the healthcare practitioner. Ina particular embodiment, the dose is about 240 mg/m².

Typically, Compound 1 Pattern 1, for example Compound 1 Pattern 1dihydrochloride, dihydrate, or a formulation produced from it isadministered to the subject prior to treatment with the chemotherapeuticagent such that its concentration reaches peak serum levels before orduring treatment with the chemotherapeutic agent, allowing for theinhibition of proliferation of immune effector cells, thus protectingthem from the harmful effects of chemotherapy. In some embodiments,Compound 1 Pattern 1, for example Compound 1 Pattern 1 dihydrochloride,dihydrate, or a formulation produced from it is administeredconcomitantly, or closely thereto, with the chemotherapeutic agentexposure. Alternatively, the CDK4/6 inhibitor described herein can beadministered following exposure to the chemotherapeutic agent if desiredto mitigate immune effector cell damage associated with chemotherapeuticagent exposure. In one embodiment, an aqueous solution produced formCompound 1 Pattern 1 is administered to a patient in need thereof.

In some embodiments, Compound 1 Pattern 1, for example Compound 1Pattern 1 dihydrochloride, dihydrate, or a formulation produced from itis administered to the subject less than about 24 hours, about 20 hours,about 16 hours, about 12 hours, about 8 hours, about 4 hours, about 2.5hours, about 2 hours, about 1 hour, about ½ hour or less prior totreatment with the chemotherapeutic agent. In a particular embodiment,Compound 1 Pattern 1, for example Compound 1 Pattern 1 dihydrochloride,dihydrate, or a formulation produced from it is administered about ½hour prior to administration of the chemotherapeutic agent.

In some embodiments, Compound 1 Pattern 1, for example Compound 1Pattern 1 dihydrochloride, dihydrate, or a formulation produced from itis administered to the subject twice before administration of thechemotherapy. For example, in some embodiments, Compound 1 Pattern 1,for example Compound 1 Pattern 1 dihydrochloride, dihydrate, or aformulation produced from it is administered between about 18 and 28hours before the administration of the chemotherapy, and then once againat less than about 4 hours, about 2.5 hours, about 2 hours, about 1hour, about ½ hour or less prior to treatment with the chemotherapeuticagent. In a particular embodiment, Compound 1 Pattern 1, for exampleCompound 1 Pattern 1 dihydrochloride, dihydrate, or a formulationproduced from it is administered between about 22 and 26 hours prior toadministration of the chemotherapeutic agent and again about ½ hour orless prior to administration of the chemotherapeutic agent.

In certain embodiments, Compound 1 Pattern 1, for example Compound 1Pattern 1 dihydrochloride, dihydrate, or a formulation produced from itis administered prior to or concomitantly with the administration of achemotherapeutic agent, wherein the chemotherapeutic agent isadministered: for example, on day 1-3 every 21 days; on days 1-3 every28 days; on day 1 every 3 weeks; on day 1, day 8, and day 15 every 28days, on day 1 and day 8 every 28 days; on days 1 and day 8 every 21days; on days 1-5 every 21 days; 1 day a week for 6-8 weeks; on days 1,22, and 43; days 1 and 2 weekly; days 1-4 and 22-25; days 1-4; days22-25, and days 43-46; and similar type chemotherapeutic regimens. Insome embodiments, Compound 1 Pattern 1, for example Compound 1 Pattern 1dihydrochloride, dihydrate, or a formulation produced from it isadministered prior to or concomitantly with at least one administrationof the chemotherapeutic agent during a chemotherapeutic treatmentregimen. In some embodiments, Compound 1 Pattern 1, for example Compound1 Pattern 1 dihydrochloride, dihydrate, or a formulation produced fromit is administered prior to or concomitantly with one or moreadministrations of the chemotherapeutic agent during a chemotherapeutictreatment regimen. In one embodiment, Compound 1 Pattern 1, for exampleCompound 1 Pattern 1 dihydrochloride, dihydrate, or a formulationproduced from it is administered prior to or concomitantly with eachadministration of the chemotherapeutic agent during a chemotherapeutictreatment regimen.

In some embodiments, Compound 1 Pattern 1, for example Compound 1Pattern 1 dihydrochloride, dihydrate, or a formulation produced from itis administered prior to or concomitantly with each administration of achemotherapeutic agent for example during a standard chemotherapeuticprotocol such as, for example, a 21-day cycle. Following cessation ofthe standard chemotherapeutic protocol, Compound 1 Pattern 1, forexample Compound 1 Pattern 1 dihydrochloride, dihydrate, or aformulation produced from it can be further administered alone in amaintenance dose. In some embodiments, Compound 1 Pattern 1, for exampleCompound 1 Pattern 1 dihydrochloride, dihydrate, or a formulationproduced from it is further administered once a week for at least 3, 4,5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 26, 52, 104 weeks, or longer.In some embodiments, Compound 1 Pattern 1, for example Compound 1Pattern 1 dihydrochloride, dihydrate, or a formulation produced from itis administered once every 21 days following the cessation of thechemotherapeutic protocol. In one embodiment, Compound 1 Pattern 1, forexample Compound 1 Pattern 1 dihydrochloride, dihydrate, or aformulation produced from it is a fast-acting, short half-life CDK4/6inhibitor.

In some embodiments, Compound 1 Pattern 1, for example Compound 1Pattern 1 dihydrochloride, dihydrate, or a formulation produced from itis administered with a chemotherapy agent in a maintenance therapytreatment regimen following cessation of the standard chemotherapeuticprotocol. Maintenance therapy can comprise either continuation of anagent given as part of the first-line or previous regimen (continuationmaintenance) or treatment with a new agent (switch maintenance).

In some embodiments, Compound 1 Pattern 1, for example Compound 1Pattern 1 dihydrochloride, dihydrate, or a formulation produced from itis further administered in a maintenance-type therapeutic regimen,wherein Compound 1 Pattern 1, for example Compound 1 Pattern 1dihydrochloride, dihydrate, or a formulation produced from it isadministered in combination with a reduced maintenance dose ofchemotherapy at a regular dosing interval for example but not limitedto, once a week, once every two weeks, once every three weeks, once amonth, once every six weeks, once every two months, once every threemonths, or once every six months following the completion of the initialchemotherapy treatment. In some embodiments, Compound 1 Pattern 1, forexample Compound 1 Pattern 1 dihydrochloride, dihydrate, or aformulation produced from it is administered with the same agent used inthe previous phase of chemotherapy treatment. In some embodiments,Compound 1 Pattern 1, for example Compound 1 Pattern 1 dihydrochloride,dihydrate, or a formulation produced from it is administered with adifferent chemotherapy agent than was used in the previous phase ofchemotherapy treatment.

Standard cancer chemotherapy can promote tumor immunity in two majorways: (i) inducing immunogenic cell death as part of its intendedtherapeutic effect; and (ii) disrupting strategies that tumors use toevade the immune response. A large body of data demonstrates that somechemotherapy drugs at their standard dose and schedule mediate theirantitumor effect, at least in part, by inducing immunogenic cell death(see, e.g., Emens et al., Chemotherapy: friend of foe to cancervaccines? Curr Opin Mol Ther 2001; 3:77-84; Vanmeerbeek et al., TrialWatch: Chemotherapy-Induced Immunogenic Cell Death in Immuni-Oncology.Oncoimmunology Vol. 9, No. 1 2020:e1703449, both incorporated byreference herein).

Immunogenic cell death (ICD) is a type of cell death characterized by,for example, cell surface translocation of calreticulin (CRT),extracellular release of ATP and high mobility group box 1 (HMBG1), andstimulation of type I interferon (IFN) responses. ICD in cancer cellsmay prime an anticancer immune response. A variety of chemotherapeuticagents can induce ICD, as indicated by the alterations intumor-infiltrating lymphocytes (TIL) abundance and composition.

In response to ICD-inducing chemotherapeutics, tumor cells expose CRT oncell surface prior to death, and release damage-associated molecularpattern (DAMP) molecules such as ATP during apoptosis or HMGB1 uponsecondary necrosis. These DAMPs stimulate the recruitment of dendriticcells (DCs) into the tumor bed, the uptake and processing of tumorantigens, and the optimal antigen presentation to T cells. Cross-primingof CD8+ T-cells is triggered by mature DCs and γδ T-cells in an IL-1βand IL-17 dependent manner. Primed CTLs then elicit a direct cytotoxicresponse to kill remaining tumor cells through the generation of IFN-γ,perforin-1 and granzyme B. In certain embodiments Compound 1 Pattern 1,for example Compound 1 Pattern 1 dihydrochloride, dihydrate, or aformulation produced from it is used for myelopreservation incombination with ICD-inducing chemotherapy.

ICD-inducing chemotherapies for use in the present invention includealkylating agents such as cyclophosphamide, trabectedin, temozolomide,melphalan, dacarbazine, and oxaliplatin; antimetabolites such asmethotrexate, mitroxantrone, gemcitabine, and 5-fluorouracil (5-FU);cytotoxic antibiotics such as bleomycin and anthracyclines, includingdoxorubicin, daunorubicin, epirubicin, idarubicin, and valrubicin;taxanes, such as paclitaxel, cabazitaxel, and docetaxel; topoisomeraseinhibitors such as topotecan, irinotecan, and etoposide; platinumcompounds such as carboplatin and cisplatin; anti-microtubule vincaalkaloid agents such as vinblastine, vincristine, vinorelbine, andvindesine. Other ICD-inducing chemotherapies include bortezomib, aninhibitor of the 26S proteasome subunit, mechlorethamine, diaziquone,mitomycin C, fludarabine and cytosine arabinoside. In some embodiments,the ICD-inducing chemotherapy is selected from idarubicin, epirubicin,doxorubicin, mitoxantrone, oxaliplatin, bortezomib, gemcitabine, andcyclophosphamide, and combinations thereof. In an alternativeembodiment, the chemotherapeutic administered is capable of inducing animmune-response may modulate tumor immunity by mechanisms distinct fromimmunogenic cell death. Various chemotherapy drugs can modulate theactivity of distinct immune cell subsets or the immune phenotype oftumor cells through enhancing antigen presentation, enhancing expressionof costimulatory molecules including B7.1 (CD80) and B7.2 (CD86),downregulating checkpoint molecules such as programmed death-ligand 1(PD-L1), or promoting tumor cell death through the fas, perforin, orGranzyme B pathways. Chemotherapies that modulate tumor immunity may doso by: abrogating myeloid-derived suppressor cell (MDSC) activity, forexample gemcitabine, 5-fluoruracil, cisplatin, and doxorubicin;abrogating Treg activity, for example cyclophosphamide, 5-fluorouracil;paclitaxel, cisplatin, and fludarabine; enhancement of T-cell crosspriming, for example gemcitabine and anthracyclines such as doxorubicin,daunorubicin, epirubicin, valrubicin and idarubicin.; augmentingdendritic cell activation, for example anthracyclines, taxanes,cyclophosphamide, vinca alkaloids, methotrexate, and mitomycin C;promoting anti-tumor CD4+ T-cell phenotype, for example cyclophosphamideand paclitaxel; and promoting tumor cell recognition and lysis, forexample cyclophosphamide, 5-fluorouracil, paclitaxel, doxorubicin,cisplatin, and cytosine arabinoside.

In some embodiments, a method for selecting a patient or patientpopulation for cancer therapy that includes the administration ofCompound 1 Pattern 1, for example Compound 1 Pattern 1 dihydrochloride,dihydrate, or a formulation produced from it with chemotherapy in amanner that increases the progression free survival or overall survivalof the patient or patient population is provided comprising, determiningif the cancer has a surrounding microenvironment that is favorable toimmune modulation, is immunogenically susceptible to CDK4/6 inhibitortreatment, or is immunogenic, and if so, administering to the patient aneffective amount of Compound 1 Pattern 1, for example Compound 1 Pattern1 dihydrochloride, dihydrate, or a formulation produced from it incombination with an effective amount of an alkylating agent such ascyclophosphamide, trabectedin, temozolomide, melphalan, dacarbazine, oroxaliplatin; an antimetabolite such as methotrexate, mitroxantrone,gemcitabine, or 5-fluorouracil (5-FU); a cytotoxic antibiotic such asbleomycin or an anthracycline such as doxorubicin, daunorubicin,epirubicin, idarubicin, or valrubicin; a taxane, such as paclitaxel,cabazitaxel, and docetaxel; topoisomerase inhibitors such as topotecan,irinotecan, and etoposide; platinum compounds such as carboplatin andcisplatin; anti-microtubule vinca alkaloid agents such as vinblastine,vincristine, vinorelbine and vindesine; bortezomib; mechlorethamine;diaziquone;; fludarabine; mitomycin C; and cytosine arabinoside. In someembodiments, the administration of Compound 1 Pattern 1, for exampleCompound 1 Pattern 1 dihydrochloride, dihydrate, or a formulationproduced from it in combination with the chemotherapy does not includeadministering an immune checkpoint inhibitor. In some embodiments, thepatient has a tumor classified as immunogenic. In some embodiments, thepatient has a hot immune tumor. In some embodiments, the patient has analtered-immunosuppressed immune tumor. In some embodiments, the patienthas an altered-excluded immune tumor. In some embodiments, the patienthas a cold tumor. In some embodiments, the patient has a tumor that isclassified as a C2 “IFN-γ Dominant” class cancer. In some embodiments,the patient has a tumor that is classified as a high “IFN-γ Signature”or a high “Expanded Immune Signature.” In some embodiments, the patienthas a tumor that is PD-L1 positive.

In some embodiments, a method for selecting a patient or patientpopulation for cancer therapy that includes the administration ofCompound 1 Pattern 1, for example Compound 1 Pattern 1 dihydrochloride,dihydrate, or a formulation produced from it with chemotherapy in amanner that increases the progression free survival or overall survivalof the patient or patient population is provided comprising, determiningif the cancer has a surrounding microenvironment that is favorable toimmune modulation, is immunogenically susceptible to CDK4/6 inhibitortreatment, or is immunogenic, and if so, administering to the patient aneffective amount of Compound 1 Pattern 1, for example Compound 1 Pattern1 dihydrochloride, dihydrate, or a formulation produced from it incombination with an effective amount of cyclophosphamide. In someembodiments, the administration of Compound 1 Pattern 1, for exampleCompound 1 Pattern 1 dihydrochloride, dihydrate, or a formulationproduced from it in combination with cyclophosphamide does not includeadministering an immune checkpoint inhibitor. In some embodiments, thepatient has a tumor classified as immunogenic. In some embodiments, thepatient has a hot immune tumor. In some embodiments, the patient has analtered-immunosuppressed immune tumor. In some embodiments, the patienthas an altered-excluded immune tumor. In some embodiments, the patienthas a cold tumor. In some embodiments, the patient has a tumor that isclassified as a C2 “IFN-γ Dominant” class cancer. In some embodiments,the patient has a tumor that is classified as a high “IFN-γ Signature”or a high “Expanded Immune Signature.” In some embodiments, the patienthas a tumor that is PD-L1 positive.

In some embodiments, a method for selecting a patient or patientpopulation for cancer therapy that includes the administration of a CDK4/6 inhibitor with chemotherapy in a manner that increases theprogression free survival or overall survival of the patient or patientpopulation is provided comprising, determining if the cancer has asurrounding microenvironment that is favorable to immune modulation, isimmunogenically susceptible to CDK4/6 inhibitor treatment, or isimmunogenic, and if so, administering to the patient an effective amountof Compound 1 Pattern 1, for example Compound 1 Pattern 1dihydrochloride, dihydrate, or a formulation produced from it incombination with an effective amount of trabectedin. In someembodiments, the administration of Compound 1 Pattern 1, for exampleCompound 1 Pattern 1 dihydrochloride, dihydrate, or a formulationproduced from it in combination with trabectedin does not includeadministering an immune checkpoint inhibitor. In some embodiments, thepatient has a tumor classified as immunogenic. In some embodiments, thepatient has a hot immune tumor. In some embodiments, the patient has analtered-immunosuppressed immune tumor. In some embodiments, the patienthas an altered-excluded immune tumor. In some embodiments, the patienthas a cold tumor. In some embodiments, the patient has a tumor that isclassified as a C2 “IFN-γ Dominant” class cancer. In some embodiments,the patient has a tumor that is classified as a high “IFN-γ Signature”or a high “Expanded Immune Signature.” In some embodiments, the patienthas a tumor that is PD-L1 positive.

In some embodiments, a method for selecting a patient or patientpopulation for cancer therapy that includes the administration ofCompound 1 Pattern 1, for example Compound 1 Pattern 1 dihydrochloride,dihydrate, or a formulation produced from it with chemotherapy in amanner that increases the progression free survival or overall survivalof the patient or patient population is provided comprising, determiningif the cancer has a surrounding microenvironment that is favorable toimmune modulation, is immunogenically susceptible to CDK4/6 inhibitortreatment, or is immunogenic, and if so, administering to the patient aneffective amount of Compound 1 Pattern 1, for example Compound 1 Pattern1 dihydrochloride, dihydrate, or a formulation produced from it incombination with an effective amount of temozolomide. In someembodiments, the administration of Compound 1 Pattern 1, for exampleCompound 1 Pattern 1 dihydrochloride, dihydrate, or a formulationproduced from it inhibitor in combination with temozolomide does notinclude administering an immune checkpoint inhibitor. In someembodiments, the patient has a tumor classified as immunogenic. In someembodiments, the patient has a hot immune tumor. In some embodiments,the patient has an altered-immunosuppressed immune tumor. In someembodiments, the patient has an altered-excluded immune tumor. In someembodiments, the patient has a cold tumor. In some embodiments, thepatient has a tumor that is classified as a C2 “IFN-γ Dominant” classcancer. In some embodiments, the patient has a tumor that is classifiedas a high “IFN-γ Signature” or a high “Expanded Immune Signature.” Insome embodiments, the patient has a tumor that is PD-L1 positive.

In some embodiments, a method for selecting a patient or patientpopulation for cancer therapy that includes the administration ofCompound 1 Pattern 1, for example Compound 1 Pattern 1 dihydrochloride,dihydrate, or a formulation produced from it r with chemotherapy in amanner that increases the progression free survival or overall survivalof the patient or patient population is provided comprising, determiningif the cancer has a surrounding microenvironment that is favorable toimmune modulation, is immunogenically susceptible to CDK4/6 inhibitortreatment, or is immunogenic, and if so, administering to the patient aneffective amount of Compound 1 Pattern 1, for example Compound 1 Pattern1 dihydrochloride, dihydrate, or a formulation produced from it incombination with an effective amount of melphalan. In some embodiments,the patient has a tumor classified as immunogenic. In some embodiments,the patient has a hot immune tumor. In some embodiments, the patient hasan altered-immunosuppressed immune tumor. In some embodiments, thepatient has an altered-excluded immune tumor. In some embodiments, thepatient has a cold tumor. In some embodiments, the patient has a tumorthat is classified as a C2 “IFN-γ Dominant” class cancer. In someembodiments, the patient has a tumor that is classified as a high “IFN-γSignature” or a high “Expanded Immune Signature.” In some embodiments,the patient has a tumor that is PD-L1 positive.

In some embodiments, a method for selecting a patient or patientpopulation for cancer therapy that includes the administration of a CDK4/6 inhibitor with chemotherapy in a manner that increases theprogression free survival or overall survival of the patient or patientpopulation is provided comprising, determining if the cancer has asurrounding microenvironment that is favorable to immune modulation, isimmunogenically susceptible to CDK4/6 inhibitor treatment, or isimmunogenic, and if so, administering to the patient an effective amountof Compound 1 Pattern 1, for example Compound 1 Pattern 1dihydrochloride, dihydrate, or a formulation produced from it incombination with an effective amount of dacarbazine. In someembodiments, the patient has a tumor classified as immunogenic. In someembodiments, the patient has a hot immune tumor. In some embodiments,the patient has an altered-immunosuppressed immune tumor. In someembodiments, the patient has an altered-excluded immune tumor. In someembodiments, the patient has a cold tumor. In some embodiments, thepatient has a tumor that is classified as a C2 “IFN-γ Dominant” classcancer. In some embodiments, the patient has a tumor that is classifiedas a high “IFN-γ Signature” or a high “Expanded Immune Signature.” Insome embodiments, the patient has a tumor that is PD-L1 positive.

In some embodiments, a method for selecting a patient or patientpopulation for cancer therapy that includes the administration ofCompound 1 Pattern 1, for example Compound 1 Pattern 1 dihydrochloride,dihydrate, or a formulation produced from it with chemotherapy in amanner that increases the progression free survival or overall survivalof the patient or patient population is provided comprising, determiningif the cancer has a surrounding microenvironment that is favorable toimmune modulation, is immunogenically susceptible to CDK4/6 inhibitortreatment, or is immunogenic, and if so, administering to the patient aneffective amount of Compound 1 Pattern 1, for example Compound 1 Pattern1 dihydrochloride, dihydrate, or a formulation produced from it incombination with an effective amount of oxaliplatin. In someembodiments, the patient has a tumor classified as immunogenic. In someembodiments, the patient has a hot immune tumor. In some embodiments,the patient has an altered-immunosuppressed immune tumor. In someembodiments, the patient has an altered-excluded immune tumor. In someembodiments, the patient has a cold tumor. In some embodiments, thepatient has a tumor that is classified as a C2 “IFN-γ Dominant” classcancer. In some embodiments, the patient has a tumor that is classifiedas a high “IFN-γ Signature” or a high “Expanded Immune Signature.” Insome embodiments, the patient has a tumor that is PD-L1 positive.

In some embodiments, a method for selecting a patient or patientpopulation for cancer therapy that includes the administration ofCompound 1 Pattern 1, for example Compound 1 Pattern 1 dihydrochloride,dihydrate, or a formulation produced from it with chemotherapy in amanner that increases the progression free survival or overall survivalof the patient or patient population is provided comprising, determiningif the cancer has a surrounding microenvironment that is favorable toimmune modulation, is immunogenically susceptible to CDK4/6 inhibitortreatment, or is immunogenic, and if so, administering to the patient aneffective amount of Compound 1 Pattern 1, for example Compound 1 Pattern1 dihydrochloride, dihydrate, or a formulation produced from it incombination with an effective amount of methotrexate. In someembodiments, the patient has a tumor classified as immunogenic. In someembodiments, the patient has a hot immune tumor. In some embodiments,the patient has an altered-immunosuppressed immune tumor. In someembodiments, the patient has an altered-excluded immune tumor. In someembodiments, the patient has a cold tumor. In some embodiments, thepatient has a tumor that is classified as a C2 “IFN-γ Dominant” classcancer. In some embodiments, the patient has a tumor that is classifiedas a high “IFN-γ Signature” or a high “Expanded Immune Signature.” Insome embodiments, the patient has a tumor that is PD-L1 positive.

In some embodiments, a method for selecting a patient or patientpopulation for cancer therapy that includes the administration of a CDK4/6 inhibitor with chemotherapy in a manner that increases theprogression free survival or overall survival of the patient or patientpopulation is provided comprising, determining if the cancer has asurrounding microenvironment that is favorable to immune modulation, isimmunogenically susceptible to CDK4/6 inhibitor treatment, or isimmunogenic, and if so, administering to the patient an effective amountof Compound 1 Pattern 1, for example Compound 1 Pattern 1dihydrochloride, dihydrate, or a formulation produced from it incombination with an effective amount of 5-fluorouracil (5-FU). In someembodiments, the patient has a tumor classified as immunogenic. In someembodiments, the patient has a hot immune tumor. In some embodiments,the patient has an altered-immunosuppressed immune tumor. In someembodiments, the patient has an altered-excluded immune tumor. In someembodiments, the patient has a cold tumor. In some embodiments, thepatient has a tumor that is classified as a C2 “IFN-γ Dominant” classcancer. In some embodiments, the patient has a tumor that is classifiedas a high “IFN-γ Signature” or a high “Expanded Immune Signature.” Insome embodiments, the patient has a tumor that is PD-L1 positive.

In some embodiments, a method for selecting a patient or patientpopulation for cancer therapy that includes the administration ofCompound 1 Pattern 1, for example Compound 1 Pattern 1 dihydrochloride,dihydrate, or a formulation produced from it with chemotherapy in amanner that increases the progression free survival or overall survivalof the patient or patient population is provided comprising, determiningif the cancer has a surrounding microenvironment that is favorable toimmune modulation, is immunogenically susceptible to CDK4/6 inhibitortreatment, or is immunogenic, and if so, administering to the patient aneffective amount of Compound 1 Pattern 1, for example Compound 1 Pattern1 dihydrochloride, dihydrate, or a formulation produced from it incombination with an effective amount of gemcitabine. In someembodiments, the patient has a tumor classified as immunogenic. In someembodiments, the patient has a hot immune tumor. In some embodiments,the patient has an altered-immunosuppressed immune tumor. In someembodiments, the patient has an altered-excluded immune tumor. In someembodiments, the patient has a cold tumor. In some embodiments, thepatient has a tumor that is classified as a C2 “IFN-γ Dominant” classcancer. In some embodiments, the patient has a tumor that is classifiedas a high “IFN-γ Signature” or a high “Expanded Immune Signature.”

In some embodiments, a method for selecting a patient or patientpopulation for cancer therapy that includes the administration ofCompound 1 Pattern 1, for example Compound 1 Pattern 1 dihydrochloride,dihydrate, or a formulation produced from it with chemotherapy in amanner that increases the progression free survival or overall survivalof the patient or patient population is provided comprising, determiningif the cancer has a surrounding microenvironment that is favorable toimmune modulation, is immunogenically susceptible to CDK4/6 inhibitortreatment, or is immunogenic, and if so, administering to the patient aneffective amount of Compound 1 Pattern 1, for example Compound 1 Pattern1 dihydrochloride, dihydrate, or a formulation produced from it incombination with an effective amount of mitoxantrone. In someembodiments, the patient has a tumor classified as immunogenic. In someembodiments, the patient has a hot immune tumor. In some embodiments,the patient has an altered-immunosuppressed immune tumor. In someembodiments, the patient has an altered-excluded immune tumor. In someembodiments, the patient has a cold tumor. In some embodiments, thepatient has a tumor that is classified as a C2 “IFN-γ Dominant” classcancer. In some embodiments, the patient has a tumor that is classifiedas a high “IFN-γ Signature” or a high “Expanded Immune Signature.” Insome embodiments, the patient has a tumor that is PD-L1 positive.

In some embodiments, a method for selecting a patient or patientpopulation for cancer therapy that includes the administration of a CDK4/6 inhibitor with chemotherapy in a manner that increases theprogression free survival or overall survival of the patient or patientpopulation is provided comprising, determining if the cancer has asurrounding microenvironment that is favorable to immune modulation, isimmunogenically susceptible to CDK4/6 inhibitor treatment, or isimmunogenic, and if so, administering to the patient an effective amountof Compound 1 Pattern 1, for example Compound 1 Pattern 1dihydrochloride, dihydrate, or a formulation produced from it incombination with an effective amount of doxorubicin. In someembodiments, the administration of Compound 1 Pattern 1, for exampleCompound 1 Pattern 1 dihydrochloride, dihydrate, or a formulationproduced from it in combination with doxorubicin does not includeadministering an immune checkpoint inhibitor. In some embodiments, thepatient has a tumor classified as immunogenic. In some embodiments, thepatient has a hot immune tumor. In some embodiments, the patient has analtered-immunosuppressed immune tumor. In some embodiments, the patienthas an altered-excluded immune tumor. In some embodiments, the patienthas a cold tumor. In some embodiments, the patient has a tumor that isclassified as a C2 “IFN-γ Dominant” class cancer. In some embodiments,the patient has a tumor that is classified as a high “IFN-γ Signature”or a high “Expanded Immune Signature.” In some embodiments, the patienthas a tumor that is PD-L1 positive.

In some embodiments, a method for selecting a patient or patientpopulation for cancer therapy that includes the administration ofCompound 1 Pattern 1, for example Compound 1 Pattern 1 dihydrochloride,dihydrate, or a formulation produced from it with chemotherapy in amanner that increases the progression free survival or overall survivalof the patient or patient population is provided comprising, determiningif the cancer has a surrounding microenvironment that is favorable toimmune modulation, is immunogenically susceptible to CDK4/6 inhibitortreatment, or is immunogenic, and if so, administering to the patient aneffective amount of Compound 1 Pattern 1, for example Compound 1 Pattern1 dihydrochloride, dihydrate, or a formulation produced from it incombination with an effective amount of daunorubicin. In someembodiments, the patient has a tumor classified as immunogenic. In someembodiments, the patient has a hot immune tumor. In some embodiments,the patient has an altered-immunosuppressed immune tumor. In someembodiments, the patient has an altered-excluded immune tumor. In someembodiments, the patient has a cold tumor. In some embodiments, thepatient has a tumor that is classified as a C2 “IFN-γ Dominant” classcancer. In some embodiments, the patient has a tumor that is classifiedas a high “IFN-γ Signature” or a high “Expanded Immune Signature.” Insome embodiments, the patient has a tumor that is PD-L1 positive.

In some embodiments, a method for selecting a patient or patientpopulation for cancer therapy that includes the administration ofCompound 1 Pattern 1, for example Compound 1 Pattern 1 dihydrochloride,dihydrate, or a formulation produced from it with chemotherapy in amanner that increases the progression free survival or overall survivalof the patient or patient population is provided comprising, determiningif the cancer has a surrounding microenvironment that is favorable toimmune modulation, is immunogenically susceptible to CDK4/6 inhibitortreatment, or is immunogenic, and if so, administering to the patient aneffective amount of Compound 1 Pattern 1, for example Compound 1 Pattern1 dihydrochloride, dihydrate, or a formulation produced from it incombination with an effective amount of idarubicin In some embodiments,the patient has a tumor classified as immunogenic. In some embodiments,the patient has a hot immune tumor. In some embodiments, the patient hasan altered-immunosuppressed immune tumor. In some embodiments, thepatient has an altered-excluded immune tumor. In some embodiments, thepatient has a cold tumor. In some embodiments, the patient has a tumorthat is classified as a C2 “IFN-γ Dominant” class cancer. In someembodiments, the patient has a tumor that is classified as a high “IFN-γSignature” or a high “Expanded Immune Signature.” In some embodiments,the patient has a tumor that is PD-L1 positive.

In some embodiments, a method for selecting a patient or patientpopulation for cancer therapy that includes the administration ofCompound 1 Pattern 1, for example Compound 1 Pattern 1 dihydrochloride,dihydrate, or a formulation produced from it with chemotherapy in amanner that increases the progression free survival or overall survivalof the patient or patient population is provided comprising, determiningif the cancer has a surrounding microenvironment that is favorable toimmune modulation, is immunogenically susceptible to CDK4/6 inhibitortreatment, or is immunogenic, and if so, administering to the patient aneffective amount of Compound 1 Pattern 1, for example Compound 1 Pattern1 dihydrochloride, dihydrate, or a formulation produced from it incombination with an effective amount of valrubicin. In some embodiments,the patient has a tumor classified as immunogenic. In some embodiments,the patient has a hot immune tumor. In some embodiments, the patient hasan altered-immunosuppressed immune tumor. In some embodiments, thepatient has an altered-excluded immune tumor. In some embodiments, thepatient has a cold tumor. In some embodiments, the patient has a tumorthat is classified as a C2 “IFN-γ Dominant” class cancer. In someembodiments, the patient has a tumor that is classified as a high “IFN-γSignature” or a high “Expanded Immune Signature.” In some embodiments,the patient has a tumor that is PD-L1 positive.

In some embodiments, a method for selecting a patient or patientpopulation for cancer therapy that includes the administration of a CDK4/6 inhibitor with chemotherapy in a manner that increases theprogression free survival or overall survival of the patient or patientpopulation is provided comprising, determining if the cancer has asurrounding microenvironment that is favorable to immune modulation, isimmunogenically susceptible to CDK4/6 inhibitor treatment, or isimmunogenic, and if so, administering to the patient an effective amountof Compound 1 Pattern 1, for example Compound 1 Pattern 1dihydrochloride, dihydrate, or a formulation produced from it incombination with an effective amount of epirubicin. In some embodiments,the patient has a tumor classified as immunogenic. In some embodiments,the patient has a hot immune tumor. In some embodiments, the patient hasan altered-immunosuppressed immune tumor. In some embodiments, thepatient has an altered-excluded immune tumor. In some embodiments, thepatient has a cold tumor. In some embodiments, the patient has a tumorthat is classified as a C2 “IFN-γ Dominant” class cancer. In someembodiments, the patient has a tumor that is classified as a high “IFN-γSignature” or a high “Expanded Immune Signature.” In some embodiments,the patient has a tumor that is PD-L1 positive.

In some embodiments, a method for selecting a patient or patientpopulation for cancer therapy that includes the administration ofCompound 1 Pattern 1, for example Compound 1 Pattern 1 dihydrochloride,dihydrate, or a formulation produced from it with chemotherapy in amanner that increases the progression free survival or overall survivalof the patient or patient population is provided comprising, determiningif the cancer has a surrounding microenvironment that is favorable toimmune modulation, is immunogenically susceptible to CDK4/6 inhibitortreatment, or is immunogenic, and if so, administering to the patient aneffective amount of Compound 1 Pattern 1, for example Compound 1 Pattern1 dihydrochloride, dihydrate, or a formulation produced from itcombination with an effective amount of bleomycin. In some embodiments,the administration of Compound 1 Pattern 1, for example Compound 1Pattern 1 dihydrochloride, dihydrate, or a formulation produced from itin combination with bleomycin does not include administering an immunecheckpoint inhibitor. In some embodiments, the patient has a hot immunetumor. In some embodiments, the patient has a tumor classified asimmunogenic. In some embodiments, the patient has a hot immune tumor. Insome embodiments, the patient has an altered-immunosuppressed immunetumor. In some embodiments, the patient has an altered-excluded immunetumor. In some embodiments, the patient has a cold tumor. In someembodiments, the patient has a tumor that is classified as a C2 “IFN-γDominant” class cancer. In some embodiments, the patient has a tumorthat is classified as a high “IFN-γ Signature” or a high “ExpandedImmune Signature.”

In some embodiments, a method for selecting a patient or patientpopulation for cancer therapy that includes the administration ofCompound 1 Pattern 1, for example Compound 1 Pattern 1 dihydrochloride,dihydrate, or a formulation produced from it with chemotherapy in amanner that increases the progression free survival or overall survivalof the patient or patient population is provided comprising, determiningif the cancer has a surrounding microenvironment that is favorable toimmune modulation, is immunogenically susceptible to CDK4/6 inhibitortreatment, or is immunogenic, and if so, administering to the patient aneffective amount of Compound 1 Pattern 1, for example Compound 1 Pattern1 dihydrochloride, dihydrate, or a formulation produced from it incombination with an effective amount of bortezomib. In some embodiments,the administration of Compound 1 Pattern 1, for example Compound 1Pattern 1 dihydrochloride, dihydrate, or a formulation produced from itin combination with bortezomib does not include administering an immunecheckpoint inhibitor. In some embodiments, the patient has a tumorclassified as immunogenic. In some embodiments, the patient has a hotimmune tumor. In some embodiments, the patient has analtered-immunosuppressed immune tumor. In some embodiments, the patienthas an altered-excluded immune tumor. In some embodiments, the patienthas a cold tumor. In some embodiments, the patient has a tumor that isclassified as a C2 “IFN-γ Dominant” class cancer. In some embodiments,the patient has a tumor that is classified as a high “IFN-γ Signature”or a high “Expanded Immune Signature.” In some embodiments, the patienthas a tumor that is PD-L1 positive.

In some embodiments, a method for selecting a patient or patientpopulation for cancer therapy that includes the administration ofCompound 1 Pattern 1, for example Compound 1 Pattern 1 dihydrochloride,dihydrate, or a formulation produced from it with chemotherapy in amanner that increases the progression free survival or overall survivalof the patient or patient population is provided comprising, determiningif the cancer has a surrounding microenvironment that is favorable toimmune modulation, is immunogenically susceptible to CDK4/6 inhibitortreatment, or is immunogenic, and if so, administering to the patient aneffective amount of Compound 1 Pattern 1, for example Compound 1 Pattern1 dihydrochloride, dihydrate, or a formulation produced from it incombination with an effective amount of paclitaxel. In some embodiments,the administration of Compound 1 Pattern 1, for example Compound 1Pattern 1 dihydrochloride, dihydrate, or a formulation produced from itin combination with paclitaxel does not include administering an immunecheckpoint inhibitor. In some embodiments, the patient has a tumorclassified as immunogenic. In some embodiments, the patient has a hotimmune tumor. In some embodiments, the patient has analtered-immunosuppressed immune tumor. In some embodiments, the patienthas an altered-excluded immune tumor. In some embodiments, the patienthas a cold tumor. In some embodiments, the patient has a tumor that isclassified as a C2 “IFN-γ Dominant” class cancer. In some embodiments,the patient has a tumor that is classified as a high “IFN-γ Signature”or a high “Expanded Immune Signature.” In some embodiments, the patienthas a tumor that is PD-L1 positive.

In some embodiments, a method for selecting a patient or patientpopulation for cancer therapy that includes the administration ofCompound 1 Pattern 1, for example Compound 1 Pattern 1 dihydrochloride,dihydrate, or a formulation produced from it with chemotherapy in amanner that increases the progression free survival or overall survivalof the patient or patient population is provided comprising, determiningif the cancer has a surrounding microenvironment that is favorable toimmune modulation, is immunogenically susceptible to CDK4/6 inhibitortreatment, or is immunogenic, and if so, administering to the patient aneffective amount of Compound 1 Pattern 1, for example Compound 1 Pattern1 dihydrochloride, dihydrate, or a formulation produced from it Compound1 Pattern 1, for example Compound 1 Pattern 1 dihydrochloride,dihydrate, or a formulation produced from it in combination with aneffective amount of docetaxel. In some embodiments, the administrationof Compound 1 Pattern 1, for example Compound 1 Pattern 1dihydrochloride, dihydrate, or a formulation produced from it incombination with docetaxel does not include administering an immunecheckpoint inhibitor. In some embodiments, the patient has a tumorclassified as immunogenic. In some embodiments, the patient has a hotimmune tumor. In some embodiments, the patient has analtered-immunosuppressed immune tumor. In some embodiments, the patienthas an altered-excluded immune tumor. In some embodiments, the patienthas a cold tumor. In some embodiments, the patient has a tumor that isclassified as a C2 “IFN-γ Dominant” class cancer. In some embodiments,the patient has a tumor that is classified as a high “IFN-γ Signature”or a high “Expanded Immune Signature.” In some embodiments, the patienthas a tumor that is PD-L1 positive.

In some embodiments, a method for selecting a patient or patientpopulation for cancer therapy that includes the administration ofCompound 1 Pattern 1, for example Compound 1 Pattern 1 dihydrochloride,dihydrate, or a formulation produced from it with chemotherapy in amanner that increases the progression free survival or overall survivalof the patient or patient population is provided comprising, determiningif the cancer has a surrounding microenvironment that is favorable toimmune modulation, is immunogenically susceptible to CDK4/6 inhibitortreatment, or is immunogenic, and if so, administering to the patient aneffective amount of Compound 1 Pattern 1, for example Compound 1 Pattern1 dihydrochloride, dihydrate, or a formulation produced from it incombination with an effective amount of cabazitaxel. In someembodiments, the administration of Compound 1 Pattern 1, for exampleCompound 1 Pattern 1 dihydrochloride, dihydrate, or a formulationproduced from it in combination with cabazitaxel does not includeadministering an immune checkpoint inhibitor. In some embodiments, thepatient has a tumor classified as immunogenic. In some embodiments, thepatient has a hot immune tumor. In some embodiments, the patient has analtered-immunosuppressed immune tumor. In some embodiments, the patienthas an altered-excluded immune tumor. In some embodiments, the patienthas a cold tumor. In some embodiments, the patient has a tumor that isclassified as a C2 “IFN-γ Dominant” class cancer. In some embodiments,the patient has a tumor that is classified as a high “IFN-γ Signature”or a high “Expanded Immune Signature.” In some embodiments, the patienthas a tumor that is PD-L1 positive.

In some embodiments, a method for selecting a patient or patientpopulation for cancer therapy that includes the administration of a CDK4/6 inhibitor with chemotherapy in a manner that increases theprogression free survival or overall survival of the patient or patientpopulation is provided comprising, determining if the cancer has asurrounding microenvironment that is favorable to immune modulation, isimmunogenically susceptible to CDK4/6 inhibitor treatment, or isimmunogenic, and if so, administering to the patient an effective amountof Compound 1 Pattern 1, for example Compound 1 Pattern 1dihydrochloride, dihydrate, or a formulation produced from it incombination with an effective amount of topotecan. In some embodiments,the administration of Compound 1 Pattern 1, for example Compound 1Pattern 1 dihydrochloride, dihydrate, or a formulation produced from itin combination with topotecan does not include administering an immunecheckpoint inhibitor. In some embodiments, the patient has a tumorclassified as immunogenic. In some embodiments, the patient has a hotimmune tumor. In some embodiments, the patient has analtered-immunosuppressed immune tumor. In some embodiments, the patienthas an altered-excluded immune tumor. In some embodiments, the patienthas a cold tumor. In some embodiments, the patient has a tumor that isclassified as a C2 “IFN-γ Dominant” class cancer. In some embodiments,the patient has a tumor that is classified as a high “IFN-γ Signature”or a high “Expanded Immune Signature.” In some embodiments, the patienthas a tumor that is PD-L1 positive.

In some embodiments, a method for selecting a patient or patientpopulation for cancer therapy that includes the administration ofCompound 1 Pattern 1, for example Compound 1 Pattern 1 dihydrochloride,dihydrate, or a formulation produced from it with chemotherapy in amanner that increases the progression free survival or overall survivalof the patient or patient population is provided comprising, determiningif the cancer has a surrounding microenvironment that is favorable toimmune modulation, is immunogenically susceptible to CDK4/6 inhibitortreatment, or is immunogenic, and if so, administering to the patient aneffective amount of Compound 1 Pattern 1, for example Compound 1 Pattern1 dihydrochloride, dihydrate, or a formulation produced from it incombination with an effective amount of etoposide. In some embodiments,the administration of Compound 1 Pattern 1, for example Compound 1Pattern 1 dihydrochloride, dihydrate, or a formulation produced from itin combination with etoposide does not include administering an immunecheckpoint inhibitor. In some embodiments, the patient has a tumorclassified as immunogenic. In some embodiments, the patient has a hotimmune tumor. In some embodiments, the patient has analtered-immunosuppressed immune tumor. In some embodiments, the patienthas an altered-excluded immune tumor. In some embodiments, the patienthas a cold tumor. In some embodiments, the patient has a tumor that isclassified as a C2 “IFN-γ Dominant” class cancer. In some embodiments,the patient has a tumor that is classified as a high “IFN-γ Signature”or a high “Expanded Immune Signature.” In some embodiments, the patienthas a tumor that is PD-L1 positive.

In some embodiments, a method for selecting a patient or patientpopulation for cancer therapy that includes the administration ofCompound 1 Pattern 1, for example Compound 1 Pattern 1 dihydrochloride,dihydrate, or a formulation produced from it with chemotherapy in amanner that increases the progression free survival or overall survivalof the patient or patient population is provided comprising, determiningif the cancer has a surrounding microenvironment that is favorable toimmune modulation, is immunogenically susceptible to CDK4/6 inhibitortreatment, or is immunogenic, and if so, administering to the patient aneffective amount of Compound 1 Pattern 1, for example Compound 1 Pattern1 dihydrochloride, dihydrate, or a formulation produced from it incombination with an effective amount of irinotecan. In some embodiments,the administration of Compound 1 Pattern 1, for example Compound 1Pattern 1 dihydrochloride, dihydrate, or a formulation produced from itin combination with irinotecan does not include administering an immunecheckpoint inhibitor. In some embodiments, the patient has a tumorclassified as immunogenic. In some embodiments, the patient has a hotimmune tumor. In some embodiments, the patient has analtered-immunosuppressed immune tumor. In some embodiments, the patienthas an altered-excluded immune tumor. In some embodiments, the patienthas a cold tumor. In some embodiments, the patient has a tumor that isclassified as a C2 “IFN-γ Dominant” class cancer. In some embodiments,the patient has a tumor that is classified as a high “IFN-γ Signature”or a high “Expanded Immune Signature.” In some embodiments, the patienthas a tumor that is PD-L1 positive.

In some embodiments, a method for selecting a patient or patientpopulation for cancer therapy that includes the administration ofCompound 1 Pattern 1, for example Compound 1 Pattern 1 dihydrochloride,dihydrate, or a formulation produced from it with chemotherapy in amanner that increases the progression free survival or overall survivalof the patient or patient population is provided comprising, determiningif the cancer has a surrounding microenvironment that is favorable toimmune modulation, is immunogenically susceptible to CDK4/6 inhibitortreatment, or is immunogenic, and if so, administering to the patient aneffective amount of Compound 1 Pattern 1, for example Compound 1 Pattern1 dihydrochloride, dihydrate, or a formulation produced from it incombination with an effective amount of cisplatin. In some embodiments,the administration of Compound 1 Pattern 1, for example Compound 1Pattern 1 dihydrochloride, dihydrate, or a formulation produced from itin combination with cisplatin does not include administering an immunecheckpoint inhibitor. In some embodiments, the patient has a tumorclassified as immunogenic. In some embodiments, the patient has a hotimmune tumor. In some embodiments, the patient has analtered-immunosuppressed immune tumor. In some embodiments, the patienthas an altered-excluded immune tumor. In some embodiments, the patienthas a cold tumor. In some embodiments, the patient has a tumor that isclassified as a C2 “IFN-γ Dominant” class cancer. In some embodiments,the patient has a tumor that is classified as a high “IFN-γ Signature”or a high “Expanded Immune Signature.” In some embodiments, the patienthas a tumor that is PD-L1 positive.

In some embodiments, a method for selecting a patient or patientpopulation for cancer therapy that includes the administration ofCompound 1 Pattern 1, for example Compound 1 Pattern 1 dihydrochloride,dihydrate, or a formulation produced from it with chemotherapy in amanner that increases the progression free survival or overall survivalof the patient or patient population is provided comprising, determiningif the cancer has a surrounding microenvironment that is favorable toimmune modulation, is immunogenically susceptible to CDK4/6 inhibitortreatment, or is immunogenic, and if so, administering to the patient aneffective amount of Compound 1 Pattern 1, for example Compound 1 Pattern1 dihydrochloride, dihydrate, or a formulation produced from it incombination with an effective amount of carboplatin. In someembodiments, the administration of Compound 1 Pattern 1, for exampleCompound 1 Pattern 1 dihydrochloride, dihydrate, or a formulationproduced from it in combination with carboplatin does not includeadministering an immune checkpoint inhibitor. In some embodiments, thepatient has a tumor classified as immunogenic. In some embodiments, thepatient has a hot immune tumor. In some embodiments, the patient has analtered-immunosuppressed immune tumor. In some embodiments, the patienthas an altered-excluded immune tumor. In some embodiments, the patienthas a cold tumor. In some embodiments, the patient has a tumor that isclassified as a C2 “IFN-γ Dominant” class cancer. In some embodiments,the patient has a tumor that is classified as a high “IFN-γ Signature”or a high “Expanded Immune Signature.” In some embodiments, the patienthas a tumor that is PD-L1 positive.

In some embodiments, a method for selecting a patient or patientpopulation for cancer therapy that includes the administration ofCompound 1 Pattern 1, for example Compound 1 Pattern 1 dihydrochloride,dihydrate, or a formulation produced from it with chemotherapy in amanner that increases the progression free survival or overall survivalof the patient or patient population is provided comprising, determiningif the cancer has a surrounding microenvironment that is favorable toimmune modulation, is immunogenically susceptible to CDK4/6 inhibitortreatment, or is immunogenic, and if so, administering to the patient aneffective amount of Compound 1 Pattern 1, for example Compound 1 Pattern1 dihydrochloride, dihydrate, or a formulation produced from it incombination with an effective amount of vinblastine. In someembodiments, the administration of Compound 1 Pattern 1, for exampleCompound 1 Pattern 1 dihydrochloride, dihydrate, or a formulationproduced from it in combination with vinblastine does not includeadministering an immune checkpoint inhibitor. In some embodiments, thepatient has a tumor classified as immunogenic. In some embodiments, thepatient has a hot immune tumor. In some embodiments, the patient has analtered-immunosuppressed immune tumor. In some embodiments, the patienthas an altered-excluded immune tumor. In some embodiments, the patienthas a cold tumor. In some embodiments, the patient has a tumor that isclassified as a C2 “IFN-γ Dominant” class cancer. In some embodiments,the patient has a tumor that is classified as a high “IFN-γ Signature”or a high “Expanded Immune Signature.” In some embodiments, the patienthas a tumor that is PD-L1 positive.

In some embodiments, a method for selecting a patient or patientpopulation for cancer therapy that includes the administration ofCompound 1 Pattern 1, for example Compound 1 Pattern 1 dihydrochloride,dihydrate, or a formulation produced from it with chemotherapy in amanner that increases the progression free survival or overall survivalof the patient or patient population is provided comprising, determiningif the cancer has a surrounding microenvironment that is favorable toimmune modulation, is immunogenically susceptible to CDK4/6 inhibitortreatment, or is immunogenic, and if so, administering to the patient aneffective amount of Compound 1 Pattern 1, for example Compound 1 Pattern1 dihydrochloride, dihydrate, or a formulation produced from it incombination with an effective amount of vincristine. In someembodiments, the administration of Compound 1 Pattern 1, for exampleCompound 1 Pattern 1 dihydrochloride, dihydrate, or a formulationproduced from it in combination with vincristine does not includeadministering an immune checkpoint inhibitor. In some embodiments, thepatient has a tumor classified as immunogenic. In some embodiments, thepatient has a hot immune tumor. In some embodiments, the patient has analtered-immunosuppressed immune tumor. In some embodiments, the patienthas an altered-excluded immune tumor. In some embodiments, the patienthas a cold tumor. In some embodiments, the patient has a tumor that isclassified as a C2 “IFN-γ Dominant” class cancer. In some embodiments,the patient has a tumor that is classified as a high “IFN-γ Signature”or a high “Expanded Immune Signature.” In some embodiments, the patienthas a tumor that is PD-L1 positive.

In some embodiments, a method for selecting a patient or patientpopulation for cancer therapy that includes the administration ofCompound 1 Pattern 1, for example Compound 1 Pattern 1 dihydrochloride,dihydrate, or a formulation produced from it with chemotherapy in amanner that increases the progression free survival or overall survivalof the patient or patient population is provided comprising, determiningif the cancer has a surrounding microenvironment that is favorable toimmune modulation, is immunogenically susceptible to CDK4/6 inhibitortreatment, or is immunogenic, and if so, administering to the patient aneffective amount of Compound 1 Pattern 1, for example Compound 1 Pattern1 dihydrochloride, dihydrate, or a formulation produced from it incombination with an effective amount of vinorelbine. In someembodiments, the administration of Compound 1 Pattern 1, for exampleCompound 1 Pattern 1 dihydrochloride, dihydrate, or a formulationproduced from it in combination with vinorelbine does not includeadministering an immune checkpoint inhibitor. In some embodiments, thepatient has a tumor classified as immunogenic. In some embodiments, thepatient has a hot immune tumor. In some embodiments, the patient has analtered-immunosuppressed immune tumor. In some embodiments, the patienthas an altered-excluded immune tumor. In some embodiments, the patienthas a cold tumor. In some embodiments, the patient has a tumor that isclassified as a C2 “IFN-γ Dominant” class cancer. In some embodiments,the patient has a tumor that is classified as a high “IFN-γ Signature”or a high “Expanded Immune Signature.” In some embodiments, the patienthas a tumor that is PD-L1 positive.

In some embodiments, a method for selecting a patient or patientpopulation for cancer therapy that includes the administration ofCompound 1 Pattern 1, for example Compound 1 Pattern 1 dihydrochloride,dihydrate, or a formulation produced from it with chemotherapy in amanner that increases the progression free survival or overall survivalof the patient or patient population is provided comprising, determiningif the cancer has a surrounding microenvironment that is favorable toimmune modulation, is immunogenically susceptible to CDK4/6 inhibitortreatment, or is immunogenic, and if so, administering to the patient aneffective amount of Compound 1 Pattern 1, for example Compound 1 Pattern1 dihydrochloride, dihydrate, or a formulation produced from it incombination with an effective amount of vindesine. In some embodiments,the administration of Compound 1 Pattern 1, for example Compound 1Pattern 1 dihydrochloride, dihydrate, or a formulation produced from itin combination with vindesine does not include administering an immunecheckpoint inhibitor. In some embodiments, the patient has a tumorclassified as immunogenic. In some embodiments, the patient has a hotimmune tumor. In some embodiments, the patient has analtered-immunosuppressed immune tumor. In some embodiments, the patienthas an altered-excluded immune tumor. In some embodiments, the patienthas a cold tumor. In some embodiments, the patient has a tumor that isclassified as a C2 “IFN-γ Dominant” class cancer. In some embodiments,the patient has a tumor that is classified as a high “IFN-γ Signature”or a high “Expanded Immune Signature.” In some embodiments, the patienthas a tumor that is PD-L1 positive.

In some embodiments, a method for selecting a patient or patientpopulation for cancer therapy that includes the administration ofCompound 1 Pattern 1, for example Compound 1 Pattern 1 dihydrochloride,dihydrate, or a formulation produced from it with chemotherapy in amanner that increases the progression free survival or overall survivalof the patient or patient population is provided comprising, determiningif the cancer has a surrounding microenvironment that is favorable toimmune modulation, is immunogenically susceptible to CDK4/6 inhibitortreatment, or is immunogenic, and if so, administering to the patient aneffective amount of Compound 1 Pattern 1, for example Compound 1 Pattern1 dihydrochloride, dihydrate, or a formulation produced from it incombination with an effective amount of diaziquone. In some embodiments,the administration of Compound 1 Pattern 1, for example Compound 1Pattern 1 dihydrochloride, dihydrate, or a formulation produced from itin combination with diaziquone does not include administering an immunecheckpoint inhibitor. In some embodiments, the patient has a tumorclassified as immunogenic. In some embodiments, the patient has a hotimmune tumor. In some embodiments, the patient has analtered-immunosuppressed immune tumor. In some embodiments, the patienthas an altered-excluded immune tumor. In some embodiments, the patienthas a cold tumor. In some embodiments, the patient has a tumor that isclassified as a C2 “IFN-γ Dominant” class cancer. In some embodiments,the patient has a tumor that is classified as a high “IFN-γ Signature”or a high “Expanded Immune Signature.” In some embodiments, the patienthas a tumor that is PD-L1 positive.

In some embodiments, a method for selecting a patient or patientpopulation for cancer therapy that includes the administration ofCompound 1 Pattern 1, for example Compound 1 Pattern 1 dihydrochloride,dihydrate, or a formulation produced from it with chemotherapy in amanner that increases the progression free survival or overall survivalof the patient or patient population is provided comprising, determiningif the cancer has a surrounding microenvironment that is favorable toimmune modulation, is immunogenically susceptible to CDK4/6 inhibitortreatment, or is immunogenic, and if so, administering to the patient aneffective amount of Compound 1 Pattern 1, for example Compound 1 Pattern1 dihydrochloride, dihydrate, or a formulation produced from it incombination with an effective amount of mechlorethamine. In someembodiments, the administration of Compound 1 Pattern 1, for exampleCompound 1 Pattern 1 dihydrochloride, dihydrate, or a formulationproduced from it in combination with mechlorethamine does not includeadministering an immune checkpoint inhibitor. In some embodiments, thepatient has a tumor classified as immunogenic. In some embodiments, thepatient has a hot immune tumor. In some embodiments, the patient has analtered-immunosuppressed immune tumor. In some embodiments, the patienthas an altered-excluded immune tumor. In some embodiments, the patienthas a cold tumor. In some embodiments, the patient has a tumor that isclassified as a C2 “IFN-γ Dominant” class cancer. In some embodiments,the patient has a tumor that is classified as a high “IFN-γ Signature”or a high “Expanded Immune Signature.” In some embodiments, the patienthas a tumor that is PD-L1 positive.

In some embodiments, a method for selecting a patient or patientpopulation for cancer therapy that includes the administration ofCompound 1 Pattern 1, for example Compound 1 Pattern 1 dihydrochloride,dihydrate, or a formulation produced from it with chemotherapy in amanner that increases the progression free survival or overall survivalof the patient or patient population is provided comprising, determiningif the cancer has a surrounding microenvironment that is favorable toimmune modulation, is immunogenically susceptible to CDK4/6 inhibitortreatment, or is immunogenic, and if so, administering to the patient aneffective amount of Compound 1 Pattern 1, for example Compound 1 Pattern1 dihydrochloride, dihydrate, or a formulation produced from it incombination with an effective amount of mitomycin C. In someembodiments, the administration of Compound 1 Pattern 1, for exampleCompound 1 Pattern 1 dihydrochloride, dihydrate, or a formulationproduced from it in combination with mitomycin C does not includeadministering an immune checkpoint inhibitor. In some embodiments, thepatient has a tumor classified as immunogenic. In some embodiments, thepatient has a hot immune tumor. In some embodiments, the patient has analtered-immunosuppressed immune tumor. In some embodiments, the patienthas an altered-excluded immune tumor. In some embodiments, the patienthas a cold tumor. In some embodiments, the patient has a tumor that isclassified as a C2 “IFN-γ Dominant” class cancer. In some embodiments,the patient has a tumor that is classified as a high “IFN-γ Signature”or a high “Expanded Immune Signature.” In some embodiments, the patienthas a tumor that is PD-L1 positive.

In some embodiments, a method for selecting a patient or patientpopulation for cancer therapy that includes the administration ofCompound 1 Pattern 1, for example Compound 1 Pattern 1 dihydrochloride,dihydrate, or a formulation produced from it with chemotherapy in amanner that increases the progression free survival or overall survivalof the patient or patient population is provided comprising, determiningif the cancer has a surrounding microenvironment that is favorable toimmune modulation, is immunogenically susceptible to CDK4/6 inhibitortreatment, or is immunogenic, and if so, administering to the patient aneffective amount of Compound 1 Pattern 1, for example Compound 1 Pattern1 dihydrochloride, dihydrate, or a formulation produced from it incombination with an effective amount of fludarabine. In someembodiments, the administration of Compound 1 Pattern 1, for exampleCompound 1 Pattern 1 dihydrochloride, dihydrate, or a formulationproduced from it in combination with fludarabine does not includeadministering an immune checkpoint inhibitor. In some embodiments, thepatient has a tumor classified as immunogenic. In some embodiments, thepatient has a hot immune tumor. In some embodiments, the patient has analtered-immunosuppressed immune tumor. In some embodiments, the patienthas an altered-excluded immune tumor. In some embodiments, the patienthas a cold tumor. In some embodiments, the patient has a tumor that isclassified as a C2 “IFN-γ Dominant” class cancer. In some embodiments,the patient has a tumor that is classified as a high “IFN-γ Signature”or a high “Expanded Immune Signature.” In some embodiments, the patienthas a tumor that is PD-L1 positive.

In some embodiments, a method for selecting a patient or patientpopulation for cancer therapy that includes the administration ofCompound 1 Pattern 1, for example Compound 1 Pattern 1 dihydrochloride,dihydrate, or a formulation produced from it with chemotherapy in amanner that increases the progression free survival or overall survivalof the patient or patient population is provided comprising, determiningif the cancer has a surrounding microenvironment that is favorable toimmune modulation, is immunogenically susceptible to CDK4/6 inhibitortreatment, or is immunogenic, and if so, administering to the patient aneffective amount of Compound 1 Pattern 1, for example Compound 1 Pattern1 dihydrochloride, dihydrate, or a formulation produced from it incombination with an effective amount of cytosine arabinoside. In someembodiments, the administration of Compound 1 Pattern 1, for exampleCompound 1 Pattern 1 dihydrochloride, dihydrate, or a formulationproduced from it in combination with cytosine arobinoside does notinclude administering an immune checkpoint inhibitor. In someembodiments, the patient has a tumor classified as immunogenic. In someembodiments, the patient has a hot immune tumor. In some embodiments,the patient has an altered-immunosuppressed immune tumor. In someembodiments, the patient has an altered-excluded immune tumor. In someembodiments, the patient has a cold tumor. In some embodiments, thepatient has a tumor that is classified as a C2 “IFN-γ Dominant” classcancer. In some embodiments, the patient has a tumor that is classifiedas a high “IFN-γ Signature” or a high “Expanded Immune Signature.” Insome embodiments, the patient has a tumor that is PD-L1 positive.

In any of the above embodiments, the patient to be treated has beendetermined to have a cancer having a surrounding microenvironment thatis favorable to immune modulation, is immunogenic, or is immunogenicallysusceptible to CDK4/6 inhibitor treatment. Accordingly, provided thecancer fits into the category as described herein, the patient may besuitable for the described treatments. In some embodiments, the cancerto be treated is selected from the group consisting of breast cancer,including but not limited to estrogen receptor (ER)-positive breastcancer and triple negative breast cancer, non-small cell lung carcinoma,head and neck squamous cell cancer, classical Hodgkin lymphoma (cHL),diffuse large B-cell lymphoma, bladder cancer, primary mediastinalB-cell lymphoma (PBMCL), urothelial carcinoma, microsatelliteinstability-high (MSI-H) solid tumors, mismatch repair deficient (dMMR)solid tumor, gastric or gastroesophageal junction (GEJ) adenocarcinoma,squamous cell carcinoma of the esophagus, cervical cancer, endometrialcancer, cholangiocarcinoma, hepatocellular carcinoma, Merkel cellcarcinoma, renal cell carcinoma, ovarian cancer, anal canal cancer,colorectal cancer, skin cutaneous melanoma and melanoma.

In some embodiments, the patient is not administered a check pointinhibitor.

In some embodiments, the patient is administered a check pointinhibitor.

(ii) Anti-Neoplastic Therapy

In addition to being used in the production of advantageous intravenoussolutions for myelopreservation, Compound 1 Pattern 1 dihydrochloride,dihydrate can be used to treat an abnormal cellular proliferationdisorder, inflammatory disorder, immune disorder, or autoimmunedisorder.

In one aspect, a method of treating a proliferative disorder in a host,including a human, is provided comprising administering isolatedCompound 1 Pattern 1, for example as a dihydrochloride, dihydrate,optionally in a pharmaceutically acceptable carrier. Non-limitingexamples of disorders include tumors, cancers, disorders related toabnormal cellular proliferation, inflammatory disorders, immunedisorders, and autoimmune disorders.

In one embodiment Compound 1 Pattern 1 is administered parentally. Thisadministration can be daily or with treatment holidays formyelopreservation.

Compound 1 Pattern 1 is useful as a therapeutic agent in a dosage formwhen administered in an effective amount to a host, including a human,to treat a tumor, cancer (solid, non-solid, diffuse, hematological,etc.), abnormal cellular proliferation, immune disorder, inflammatorydisorder, blood disorder, a myelo- or lymphoproliferative disorder suchas B- or T-cell lymphomas, multiple myeloma, breast cancer, prostatecancer, AML, ALL, ACL, lung cancer, pancreatic cancer, colon cancer,skin cancer, melanoma, Waldenstrom's macroglobulinemia, Wiskott-Aldrichsyndrome, or a post-transplant lymphoproliferative disorder; anautoimmune disorder, for example, Lupus, Crohn' s Disease, Addisondisease, Celiac disease, dermatomyositis, Graves disease, thyroiditis,multiple sclerosis, pernicious anemia, reactive arthritis, or type Idiabetes; a disease of cardiologic malfunction, includinghypercholesterolemia; an infectious disease, including a viral and/orbacterial infection; an inflammatory condition, including asthma,chronic peptic ulcers, tuberculosis, rheumatoid arthritis,periodontitis, ulcerative colitis, or hepatitis.

Exemplary proliferative disorders include, but are not limited to,benign growths, neoplasms, tumors, cancer (Rb positive or Rb negative),autoimmune disorders, inflammatory disorders graft-versus-hostrejection, and fibrotic disorders.

Non-limiting examples of cancers that can be treated according to thepresent invention include, but are not limited to, acoustic neuroma,adenocarcinoma, adrenal gland cancer, anal cancer, angiosarcoma (e.g.,lymphangiosarcoma, lymphangioendotheliosarcoma, hemangiosarcoma),appendix cancer, benign monoclonal gammopathy, biliary cancer (e.g.,cholangiocarcinoma), bladder cancer, breast cancer (e.g., adenocarcinomaof the breast, papillary carcinoma of the breast, mammary cancer,medullary carcinoma of the breast), brain cancer (e.g., meningioma;glioma, e.g., astrocytoma, oligodendroglioma; medulloblastoma), bronchuscancer, carcinoid tumor, cervical cancer (e.g., cervicaladenocarcinoma), choriocarcinoma, chordoma, craniopharyngioma,colorectal cancer (e.g., colon cancer, rectal cancer, colorectaladenocarcinoma), epithelial carcinoma, ependymoma, endotheliosarcoma(e.g., Kaposi's sarcoma, multiple idiopathic hemorrhagic sarcoma),endometrial cancer (e.g., uterine cancer, uterine sarcoma), esophagealcancer (e.g., adenocarcinoma of the esophagus, Barrett's adenocarinoma),Ewing's sarcoma, eye cancer (e.g., intraocular melanoma,retinoblastoma), familiar hypereosinophilia, gall bladder cancer,gastric cancer (e.g., stomach adenocarcinoma), gastrointestinal stromaltumor (GIST), head and neck cancer (e.g., head and neck squamous cellcarcinoma, oral cancer (e.g., oral squamous cell carcinoma (OSCC),throat cancer (e.g., laryngeal cancer, pharyngeal cancer, nasopharyngealcancer, oropharyngeal cancer)), hematopoietic cancers (e.g., leukemiasuch as acute lymphocytic leukemia (ALL)—also known as acutelymphoblastic leukemia or acute lymphoid leukemia (e.g., B-cell ALL,T-cell ALL), acute myelocytic leukemia (AML) (e.g., B-cell AML, T-cellAML), chronic myelocytic leukemia (CML) (e.g., B-cell CML, T-cell CML),and chronic lymphocytic leukemia (CLL) (e.g., B-cell CLL, T-cell CLL);lymphoma such as Hodgkin lymphoma (HL) (e.g., B-cell HL, T-cell HL) andnon-Hodgkin lymphoma (NHL) (e.g., B-cell NHL such as diffuse large celllymphoma (DLCL) (e.g., diffuse large B-cell lymphoma (DLBCL)),follicular lymphoma, chronic lymphocytic leukemia/small lymphocyticlymphoma (CLL/SLL), mantle cell lymphoma (MCL), marginal zone B-celllymphomas (e.g., mucosa-associated lymphoid tissue (MALT) lymphomas,nodal marginal zone B-cell lymphoma, splenic marginal zone B-celllymphoma), primary mediastinal B-cell lymphoma, Burkitt lymphoma,lymphoplasmacytic lymphoma (i.e., “Waldenstrom's macroglobulinemia”),hairy cell leukemia (HCL), immunoblastic large cell lymphoma, precursorB-lymphoblastic lymphoma and primary central nervous system (CNS)lymphoma; and T-cell NHL such as precursor T-lymphoblasticlymphoma/leukemia, peripheral T-cell lymphoma (PTCL) (e.g., cutaneousT-cell lymphoma (CTCL) (e.g., mycosis fungiodes, Sezary syndrome),angioimmunoblastic T-cell lymphoma, extranodal natural killer T-celllymphoma, enteropathy type T-cell lymphoma, subcutaneouspanniculitis-like T-cell lymphoma, anaplastic large cell lymphoma); amixture of one or more leukemia/lymphoma as described above; andmultiple myeloma (MM)), heavy chain disease (e.g., alpha chain disease,gamma chain disease, mu chain disease), hemangioblastoma, inflammatorymyofibroblastic tumors, immunocytic amyloidosis, kidney cancer (e.g.,nephroblastoma a.k.a. Wilms' tumor, renal cell carcinoma), liver cancer(e.g., hepatocellular cancer (HCC), malignant hepatoma), lung cancer(e.g., bronchogenic carcinoma, small cell lung cancer (SCLC), non-smallcell lung cancer (NSCLC), adenocarcinoma of the lung), leiomyosarcoma(LMS), mastocytosis (e.g., systemic mastocytosis), myelodysplasticsyndrome (MDS), mesothelioma, myeloproliferative disorder (MPD) (e.g.,polycythemia Vera (PV), essential thrombocytosis (ET), agnogenic myeloidmetaplasia (AMM) a.k.a. myelofibrosis (MF), chronic idiopathicmyelofibrosis, chronic myelocytic leukemia (CML), chronic neutrophilicleukemia (CNL), hypereosinophilic syndrome (HES)), neuroblastoma,neurofibroma (e.g., neurofibromatosis (NF) type 1 or type 2,schwannomatosis), neuroendocrine cancer (e.g., gastroenteropancreaticneuroendoctrine tumor (GEP-NET), carcinoid tumor), osteosarcoma, ovariancancer (e.g., cystadenocarcinoma, ovarian embryonal carcinoma, ovarianadenocarcinoma), papillary adenocarcinoma, pancreatic cancer (e.g.,pancreatic andenocarcinoma, intraductal papillary mucinous neoplasm(IPMN), Islet cell tumors), penile cancer (e.g., Paget's disease of thepenis and scrotum), pinealoma, primitive neuroectodermal tumor (PNT),prostate cancer (e.g., prostate adenocarcinoma), rectal cancer,rhabdomyosarcoma, salivary gland cancer, skin cancer (e.g., squamouscell carcinoma (SCC), keratoacanthoma (KA), melanoma, basal cellcarcinoma (BCC)), small bowel cancer (e.g., appendix cancer), softtissue sarcoma (e.g., malignant fibrous histiocytoma (MFH), liposarcoma,malignant peripheral nerve sheath tumor (MPNST), chondrosarcoma,fibrosarcoma, myxosarcoma), sebaceous gland carcinoma, sweat glandcarcinoma, synovioma, testicular cancer (e.g., seminoma, testicularembryonal carcinoma), thyroid cancer (e.g., papillary carcinoma of thethyroid, papillary thyroid carcinoma (PTC), medullary thyroid cancer),urethral cancer, vaginal cancer and vulvar cancer (e.g., Paget's diseaseof the vulva).

In certain aspects, the invention includes the use of an effectiveamount of the isolated Compound 1 Pattern 1, or its pharmaceuticallyacceptable salt, prodrug or isotopic variant optionally in apharmaceutical composition, to treat a host, typically a human, with aselected cancer, tumor, hyperproliferative condition or an inflammatoryor immune disorder. Compound 1 Pattern 1 is also active against T-cellproliferation. Given the paucity of drugs for T-cell cancers andabnormal proliferation, the identification of such uses represents asubstantial improvement in the medical therapy for these diseases.

In another embodiment, the disorder is myelodysplastic syndrome (MDS).

In certain embodiments, the cancer is a hematopoietic cancer. In certainembodiments, the hematopoietic cancer is a lymphoma. In certainembodiments, the hematopoietic cancer is a leukemia. In certainembodiments, the leukemia is acute myelocytic leukemia (AML).

In certain embodiments, the proliferative disorder is amyeloproliferative neoplasm. In certain embodiments, themyeloproliferative neoplasm (MPN) is primary myelofibrosis (PMF).

In certain embodiments, the cancer is a solid tumor. A solid tumor, asused herein, refers to an abnormal mass of tissue that usually does notcontain cysts or liquid areas. Different types of solid tumors are namedfor the type of cells that form them. Examples of classes of solidtumors include, but are not limited to, sarcomas, carcinomas, andlymphomas, as described above herein. Additional examples of solidtumors include, but are not limited to, squamous cell carcinoma, coloncancer, breast cancer, prostate cancer, lung cancer, liver cancer,pancreatic cancer, and melanoma.

In certain embodiments, the condition treated with Compound 1 Pattern 1is a disorder related to abnormal cellular proliferation.

Abnormal cellular proliferation, notably hyperproliferation, can occuras a result of a wide variety of factors, including genetic mutation,infection, exposure to toxins, autoimmune disorders, and benign ormalignant tumor induction.

There are a number of skin disorders associated with cellularhyperproliferation. Psoriasis, for example, is a benign disease of humanskin generally characterized by plaques covered by thickened scales. Thedisease is caused by increased proliferation of epidermal cells ofunknown cause. Chronic eczema is also associated with significanthyperproliferation of the epidermis. Other diseases caused byhyperproliferation of skin cells include atopic dermatitis, lichenplanus, warts, pemphigus vulgaris, actinic keratosis, basal cellcarcinoma and squamous cell carcinoma.

Other hyperproliferative cell disorders include blood vesselproliferation disorders, fibrotic disorders, autoimmune disorders,graft-versus-host rejection, tumors and cancers.

Blood vessel proliferative disorders include angiogenic and vasculogenicdisorders. Proliferation of smooth muscle cells in the course ofdevelopment of plaques in vascular tissue cause, for example,restenosis, retinopathies and atherosclerosis. Both cell migration andcell proliferation play a role in the formation of atheroscleroticlesions.

Fibrotic disorders are often due to the abnormal formation of anextracellular matrix. Examples of fibrotic disorders include hepaticcirrhosis and mesangial proliferative cell disorders. Hepatic cirrhosisis characterized by the increase in extracellular matrix constituentsresulting in the formation of a hepatic scar. Hepatic cirrhosis cancause diseases such as cirrhosis of the liver. An increasedextracellular matrix resulting in a hepatic scar can also be caused byviral infection such as hepatitis. Lipocytes appear to play a major rolein hepatic cirrhosis.

Mesangial disorders are brought about by abnormal proliferation ofmesangial cells. Mesangial hyperproliferative cell disorders includevarious human renal diseases, such as glomerulonephritis, diabeticnephropathy, malignant nephrosclerosis, thrombotic micro-angiopathysyndromes, transplant rejection, and glomerulopathies.

Another disease with a proliferative component is rheumatoid arthritis.Rheumatoid arthritis is generally considered an autoimmune disease thatis thought to be associated with activity of autoreactive T cells, andto be caused by autoantibodies produced against collagen and IgE.

Other disorders that can include an abnormal cellular proliferativecomponent include Bechet's syndrome, acute respiratory distress syndrome(ARDS), ischemic heart disease, post-dialysis syndrome, leukemia,acquired immune deficiency syndrome, vasculitis, lipid histiocytosis,septic shock and inflammation in general.

In certain embodiments a compound of the present invention and itspharmaceutically acceptable derivatives or pharmaceutically acceptableformulations containing these compounds are also useful in theprevention and treatment of HBV infections and other related conditionssuch as anti-HBV antibody positive and HBV-positive conditions, chronicliver inflammation caused by HBV, cirrhosis, acute hepatitis, fulminanthepatitis, chronic persistent hepatitis, and fatigue. These compounds orformulations can also be used prophylactically to prevent or retard theprogression of clinical illness in individuals who are anti-HBV antibodyor HBV-antigen positive or who have been exposed to HBV.

In certain embodiments, the condition is associated with an immuneresponse.

Cutaneous contact hypersensitivity and asthma are just two examples ofimmune responses that can be associated with significant morbidity.Others include atopic dermatitis, eczema, Sjogren's Syndrome, includingkeratoconjunctivitis sicca secondary to Sjogren's Syndrome, alopeciaareata, allergic responses due to arthropod bite reactions, Crohn'sdisease, aphthous ulcer, iritis, conjunctivitis, keratoconjunctivitis,ulcerative colitis, cutaneous lupus erythematosus, scleroderma,vaginitis, proctitis, and drug eruptions. These conditions may result inany one or more of the following symptoms or signs: itching, swelling,redness, blisters, crusting, ulceration, pain, scaling, cracking, hairloss, scarring, or oozing of fluid involving the skin, eye, or mucosalmembranes.

In atopic dermatitis, and eczema in general, immunologically mediatedleukocyte infiltration (particularly infiltration of mononuclear cells,lymphocytes, neutrophils, and eosinophils) into the skin importantlycontributes to the pathogenesis of these diseases. Chronic eczema alsois associated with significant hyperproliferation of the epidermis.Immunologically mediated leukocyte infiltration also occurs at sitesother than the skin, such as in the airways in asthma and in the tearproducing gland of the eye in keratoconjunctivitis sicca.

In one non-limiting embodiment compounds of the present invention areused as topical agents in treating contact dermatitis, atopicdermatitis, eczematous dermatitis, psoriasis, Sjogren's Syndrome,including keratoconjunctivitis sicca secondary to Sjogren's Syndrome,alopecia areata, allergic responses due to arthropod bite reactions,Crohn's disease, aphthous ulcer, iritis, conjunctivitis,keratoconjunctivitis, ulcerative colitis, asthma, allergic asthma,cutaneous lupus erythematosus, scleroderma, vaginitis, proctitis, anddrug eruptions. The novel method may also be useful in reducing theinfiltration of skin by malignant leukocytes in diseases such as mycosisfungoides. These compounds can also be used to treat anaqueous-deficient dry eye state (such as immune mediatedkeratoconjunctivitis) in a patient suffering therefrom, by administeringthe compound topically to the eye.

The term “neoplasia” or “cancer” is used throughout the specification torefer to the pathological process that results in the formation andgrowth of a cancerous or malignant neoplasm, i.e., abnormal tissue(solid) or cells (non-solid) that grow by cellular proliferation, oftenmore rapidly than normal and continues to grow after the stimuli thatinitiated the new growth cease. Malignant neoplasms show partial orcomplete lack of structural organization and functional coordinationwith the normal tissue and most invade surrounding tissues, canmetastasize to several sites, are likely to recur after attemptedremoval and may cause the death of the patient unless adequatelytreated. As used herein, the term neoplasia is used to describe allcancerous disease states and embraces or encompasses the pathologicalprocess associated with malignant hematogenous, ascitic and solidtumors. Exemplary cancers which may be treated by the present disclosedcompounds either alone or in combination with at least one additionalanti-cancer agent include squamous-cell carcinoma, basal cell carcinoma,adenocarcinoma, hepatocellular carcinomas, and renal cell carcinomas,cancer of the bladder, bowel, breast, cervix, colon, esophagus, head,kidney, liver, lung, neck, ovary, pancreas, prostate, and stomach;leukemias; benign and malignant lymphomas, particularly Burkitt'slymphoma and Non-Hodgkin's lymphoma; benign and malignant melanomas;myeloproliferative diseases; sarcomas, including Ewing's sarcoma,hemangiosarcoma, Kaposi's sarcoma, liposarcoma, myosarcomas, peripheralneuroepithelioma, synovial sarcoma, gliomas, astrocytomas,oligodendrogliomas, ependymomas, gliobastomas, neuroblastomas,ganglioneuromas, gangliogliomas, medulloblastomas, pineal cell tumors,meningiomas, meningeal sarcomas, neurofibromas, and Schwannomas; bowelcancer, breast cancer, prostate cancer, cervical cancer, uterine cancer,lung cancer, ovarian cancer, testicular cancer, thyroid cancer,astrocytoma, esophageal cancer, pancreatic cancer, stomach cancer, livercancer, colon cancer, melanoma; carcinosarcoma, Hodgkin's disease,Wilms' tumor and teratocarcinomas. Additional cancers which may betreated using the disclosed compounds according to the present inventioninclude, for example, acute granulocytic leukemia, acute lymphocyticleukemia (ALL), acute myelogenous leukemia (AML), adenocarcinoma,adenosarcoma, adrenal cancer, adrenocortical carcinoma, anal cancer,anaplastic astrocytoma, angiosarcoma, appendix cancer, astrocytoma,Basal cell carcinoma, B-Cell lymphoma, bile duct cancer, bladder cancer,bone cancer, bone marrow cancer, bowel cancer, brain cancer, brain stemglioma, breast cancer, triple (estrogen, progesterone and HER-2)negative breast cancer, double negative breast cancer (two of estrogen,progesterone and HER-2 are negative), single negative (one of estrogen,progesterone and HER-2 is negative), estrogen-receptor positive,HER2-negative breast cancer, estrogen receptor-negative breast cancer,estrogen receptor positive breast cancer, metastatic breast cancer,luminal A breast cancer, luminal B breast cancer, Her2-negative breastcancer, HER2-positive or negative breast cancer, progesteronereceptor-negative breast cancer, progesterone receptor-positive breastcancer, recurrent breast cancer, carcinoid tumors, cervical cancer,cholangiocarcinoma, chondrosarcoma, chronic lymphocytic leukemia (CLL),chronic myelogenous leukemia (CML), colon cancer, colorectal cancer,craniopharyngioma, cutaneous lymphoma, cutaneous melanoma, diffuseastrocytoma, ductal carcinoma in situ (DCIS), endometrial cancer,ependymoma, epithelioid sarcoma, esophageal cancer, ewing sarcoma,extrahepatic bile duct cancer, eye cancer, fallopian tube cancer,fibrosarcoma, gallbladder cancer, gastric cancer, gastrointestinalcancer, gastrointestinal carcinoid cancer, gastrointestinal stromaltumors (GIST), germ cell tumor glioblastoma multiforme (GBM), glioma,hairy cell leukemia, head and neck cancer, hemangioendothelioma, Hodgkinlymphoma, hypopharyngeal cancer, infiltrating ductal carcinoma (IDC),infiltrating lobular carcinoma (ILC), inflammatory breast cancer (IBC),intestinal Cancer, intrahepatic bile duct cancer, invasive/infiltratingbreast cancer, Islet cell cancer, jaw cancer, Kaposi sarcoma, kidneycancer, laryngeal cancer, leiomyosarcoma, leptomeningeal metastases,leukemia, lip cancer, liposarcoma, liver cancer, lobular carcinoma insitu, low-grade astrocytoma, lung cancer, lymph node cancer, lymphoma,male breast cancer, medullary carcinoma, medulloblastoma, melanoma,meningioma, Merkel cell carcinoma, mesenchymal chondrosarcoma,mesenchymous, mesothelioma metastatic breast cancer, metastatic melanomametastatic squamous neck cancer, mixed gliomas, monodermal teratoma,mouth cancer mucinous carcinoma, mucosal melanoma, multiple myeloma,Mycosis Fungoides, myelodysplastic syndrome, nasal cavity cancer,nasopharyngeal cancer, neck cancer, neuroblastoma, neuroendocrine tumors(NETs), non-Hodgkin's lymphoma, non-small cell lung cancer (NSCLC), oatcell cancer, ocular cancer, ocular melanoma, oligodendroglioma, oralcancer, oral cavity cancer, oropharyngeal cancer, osteogenic sarcoma,osteosarcoma, ovarian cancer, ovarian epithelial cancer ovarian germcell tumor, ovarian primary peritoneal carcinoma, ovarian sex cordstromal tumor, Paget's disease, pancreatic cancer, papillary carcinoma,paranasal sinus cancer, parathyroid cancer, pelvic cancer, penilecancer, peripheral nerve cancer, peritoneal cancer, pharyngeal cancer,pheochromocytoma, pilocytic astrocytoma, pineal region tumor,pineoblastoma, pituitary gland cancer, primary central nervous system(CNS) lymphoma, prostate cancer, rectal cancer, renal cell carcinoma,renal pelvis cancer, rhabdomyosarcoma, salivary gland cancer, softtissue sarcoma, bone sarcoma, sarcoma, sinus cancer, skin cancer, smallcell lung cancer (SCLC), small intestine cancer, spinal cancer, spinalcolumn cancer, spinal cord cancer, squamous cell carcinoma, stomachcancer, synovial sarcoma, T-cell lymphoma, testicular cancer, throatcancer, thymoma/thymic carcinoma, thyroid cancer, tongue cancer, tonsilcancer, transitional cell cancer, tubal cancer, tubular carcinoma,undiagnosed cancer, ureteral cancer, urethral cancer, uterineadenocarcinoma, uterine cancer, uterine sarcoma, vaginal cancer, vulvarcancer, T-cell lineage acute lymphoblastic leukemia (T-ALL), T-celllineage lymphoblastic lymphoma (T-LL), peripheral T-cell lymphoma, AdultT-cell leukemia, Pre-B ALL, Pre-B lymphomas, large B-cell lymphoma,Burkitts lymphoma, B-cell ALL, Philadelphia chromosome positive ALL,Philadelphia chromosome positive CML, juvenile myelomonocytic leukemia(JMML), acute promyelocytic leukemia (a subtype of AML), large granularlymphocytic leukemia, Adult T-cell chronic leukemia, diffuse large Bcell lymphoma, follicular lymphoma; Mucosa-Associated Lymphatic Tissuelymphoma (MALT), small cell lymphocytic lymphoma, mediastinal large Bcell lymphoma, nodal marginal zone B cell lymphoma (NMZL); splenicmarginal zone lymphoma (SMZL); intravascular large B-cell lymphoma;primary effusion lymphoma; or lymphomatoid granulomatosis;; B-cellprolymphocytic leukemia; splenic lymphoma/leukemia, unclassifiable,splenic diffuse red pulp small B-cell lymphoma; lymphoplasmacyticlymphoma; heavy chain diseases, for example, Alpha heavy chain disease,Gamma heavy chain disease, Mu heavy chain disease, plasma cell myeloma,solitary plasmacytoma of bone; extraosseous plasmacytoma; primarycutaneous follicle center lymphoma, T cell/histocyte rich large B-celllymphoma, DLBCL associated with chronic inflammation; Epstein-Barr virus(EBV)+ DLBCL of the elderly; primary mediastinal (thymic) large B-celllymphoma, primary cutaneous DLBCL, leg type, ALK+ large B-cell lymphoma,plasmablastic lymphoma; large B-cell lymphoma arising in HHV8-associatedmulticentric, Castleman disease; B-cell lymphoma, unclassifiable, withfeatures intermediate between diffuse large B-cell lymphoma, or B-celllymphoma, unclassifiable, with features intermediate between diffuselarge B-cell lymphoma and classical Hodgkin lymphoma.

In another aspect, a method of increasing BIM expression (e.g., BCLC2L11expression) is provided to induce apoptosis in a cell comprisingcontacting a compound of the present invention or a pharmaceuticallyacceptable composition, salt, isotopic analog, or prodrug thereof withthe cell. In certain embodiments, the method is an in vitro method. Incertain embodiments, the method is an in vivo method. BCL2L11 expressionis tightly regulated in a cell. BCL2L11 encodes for BIM, a proapoptoticprotein. BCL2L11 is downregulated in many cancers and BIM is inhibitedin many cancers, including chronic myelocytic leukemia (CML) andnon-small cell lung cancer (NSCLC) and that suppression of BCL2L11expression can confer resistance to tyrosine kinase inhibitors. See,e.g., Ng et al., Nat. Med. (2012) 18:521-528.

In yet another aspect, a method of treating a condition associated withangiogenesis is provided, such as, for example, a diabetic condition(e.g., diabetic retinopathy), an inflammatory condition (e.g.,rheumatoid arthritis), macular degeneration, obesity, atherosclerosis,or a proliferative disorder, comprising administering to a subject inneed thereof a compound of the present invention or a pharmaceuticallyacceptable composition, salt, isotopic analog, or prodrug thereof.

In certain embodiments, the condition associated with angiogenesis ismacular degeneration. In certain embodiments, provided is a method oftreating macular degeneration comprising administering to a subject inneed thereof a compound of the present invention or a pharmaceuticallyacceptable composition, salt, isotopic analog, or prodrug thereof.

In certain embodiments, the condition associated with angiogenesis isobesity. As used herein, “obesity” and “obese” as used herein, refers toclass I obesity, class II obesity, class III obesity and pre-obesity(e.g., being “over-weight”) as defined by the World Health Organization.In certain embodiments, a method of treating obesity is providedcomprising administering to a subject in need thereof a compound of thepresent invention or a pharmaceutically acceptable composition, salt,isotopic analog, or prodrug thereof.

In certain embodiments, the condition associated with angiogenesis isatherosclerosis. In certain embodiments, provided is a method oftreating atherosclerosis comprising administering to a subject in needthereof a compound of the present invention or a pharmaceuticallyacceptable composition, salt, isotopic analog, or prodrug thereof.

In certain embodiments, the condition associated with angiogenesis is aproliferative disorder. In certain embodiments, provided is a method oftreating a proliferative disorder comprising administering to a subjectin need thereof a compound of the present invention or apharmaceutically acceptable composition, salt, isotopic analog, orprodrug thereof.

In an alternative embodiment the compound is administered at a dose ofabout 50 mg/m² to about 800 mg/m², from about 100 mg/m² to about 600mg/m², from about 100 mg/m² to about 500 mg/m², from about 100 mg/m² toabout 400 mg/m², from about 100 mg/m² to about 350 mg/m², from about 150mg/m² to about 350 mg/m², from about 200 mg/m² to about 350 mg/m², orfrom about 200 mg/m² to about 300 mg/m².

Isolated Compound 1 Pattern 1 can be used in an effective amount aloneor in combination with another compound of the present invention oranother bioactive agent to treat a host such as a human with a disorderas described herein.

The isolated Compound 1 Pattern 1 described herein can be used in aneffective amount alone or in combination with another compound of thepresent invention or another bioactive agent to treat a host such as ahuman with a disorder as described herein.

The term “bioactive agent” is used to describe an agent, other than theselected compound according to the present invention, which can be usedin combination or alternation with a compound of the present inventionto achieve a desired result of therapy. In one embodiment, the compoundof the present invention and the bioactive agent are administered in amanner that they are active in vivo during overlapping time periods, forexample, have time-period overlapping Cmax, Tmax, AUC or otherpharmacokinetic parameter. In another embodiment, isolated Compound 1Pattern 1 and the bioactive agent are administered to a host in needthereof that do not have overlapping pharmacokinetic parameter, however,one has a therapeutic impact on the therapeutic efficacy of the other.

In one aspect of this embodiment, the bioactive agent is an immunemodulator, including but not limited to a checkpoint inhibitor,including as non-limiting examples, a PD-1 inhibitor, PD-L1 inhibitor,PD-L2 inhibitor, CTLA-4 inhibitor, LAG-3 inhibitor, TIM-3 inhibitor,V-domain Ig suppressor of T-cell activation (VISTA) inhibitors, smallmolecule, peptide, nucleotide, or other inhibitor. In certain aspects,the immune modulator is an antibody, such as a monoclonal antibody.

PD-1 inhibitors that blocks the interaction of PD-1 and PD-L1 by bindingto the PD-1 receptor, and in turn inhibit immune suppression include,for example, nivolumab (Opdivo), pembrolizumab (Keytruda), pidilizumab,AMP-224 (AstraZeneca and Medlmmune), PF-06801591 (Pfizer), MEDI0680(AstraZeneca), PDR001 (Novartis), REGN2810 (Regeneron), SHR-12-1(Jiangsu Hengrui Medicine Company and Incyte Corporation), TSR-042(Tesaro), and the PD-L1/VISTA inhibitor CA-170 (Curis Inc.). PD-L1inhibitors that block the interaction of PD-1 and PD-L1 by binding tothe PD-L1 receptor, and in turn inhibits immune suppression, include forexample, atezolizumab (Tecentriq), durvalumab (AstraZeneca andMedImmune), KN035 (Alphamab), and BMS-936559 (Bristol-Myers Squibb).CTLA-4 checkpoint inhibitors that bind to CTLA-4 and inhibits immunesuppression include, but are not limited to, ipilimumab, tremelimumab(AstraZeneca and MedImmune), AGEN1884 and AGEN2041 (Agenus). LAG-3checkpoint inhibitors, include, but are not limited to, BMS-986016(Bristol-Myers Squibb), GSK2831781 (GlaxoSmithKline), IMP321 (PrimaBioMed), LAG525 (Novartis), and the dual PD-1 and LAG-3 inhibitor MGD013(MacroGenics). An example of a TIM-3 inhibitor is TSR-022 (Tesaro).

In yet another embodiment, isolated Compound 1 Pattern 1 as describedherein can be administered in an effective amount for the treatment ofabnormal tissue of the female reproductive system such as breast,ovarian, endometrial, or uterine cancer, in combination or alternationwith an effective amount of an estrogen inhibitor including but notlimited to a SERM (selective estrogen receptor modulator), a SERD(selective estrogen receptor degrader), a complete estrogen receptordegrader, or another form of partial or complete estrogen antagonist oragonist. Partial anti-estrogens like raloxifene and tamoxifen retainsome estrogen-like effects, including an estrogen-like stimulation ofuterine growth, and also, in some cases, an estrogen-like action duringbreast cancer progression which actually stimulates tumor growth. Incontrast, fulvestrant, a complete anti-estrogen, is limitedestrogen-like action on the uterus and is effective intamoxifen-resistant tumors. Non-limiting examples of anti-estrogencompounds are provided in WO 2014/19176 assigned to Astra Zeneca,WO2013/090921, WO 2014/203129, WO 2014/203132, and US2013/0178445assigned to Olema Pharmaceuticals, and U.S. Pat. Nos. 9,078,871,8,853,423, and 8,703, 810, as well as US 2015/0005286, WO 2014/205136,and WO 2014/205138.

Additional non-limiting examples of anti-estrogen compounds include:SERMS such as anordrin, bazedoxifene, broparestriol, chlorotrianisene,clomiphene citrate, cyclofenil, lasofoxifene, ormeloxifene, raloxifene,tamoxifen, toremifene, and fulvestrant; aromatase inhibitors such asaminoglutethimide, testolactone, anastrozole, exemestane, fadrozole,formestane, and letrozole; and antigonadotropins such as leuprorelin,cetrorelix, allylestrenol, chloromadinone acetate, cyproterone acetate,delmadinone acetate, dydrogesterone, medroxyprogesterone acetate,megestrol acetate, nomegestrol acetate, norethisterone acetate,progesterone, and spironolactone. Other estrogenic ligands that can beused according to the present invention are described in U.S. Pat. Nos.4,418,068; 5,478,847; 5,393,763; and 5,457,117, WO2011/156518, U.S. Pat.Nos. 8,455,534 and 8,299,112, 9,078,871; 8,853,423; 8,703,810; US2015/0005286; and WO 2014/205138, US2016/0175289, US2015/0258080, WO2014/191726, WO 2012/084711; WO 2002/013802; WO 2002/004418; WO2002/003992; WO 2002/003991; WO 2002/003990; WO 2002/003989; WO2002/003988; WO 2002/003986; WO 2002/003977; WO 2002/003976; WO2002/003975; WO 2006/078834; U.S. Pat. No. 6,821,989; US 2002/0128276;U.S. Pat. No. 6,777,424; US 2002/0016340; U.S. Pat. Nos. 6,326,392;6,756,401; US 2002/0013327; U.S. Pat. Nos. 6,512,002; 6,632,834; US2001/0056099; U.S. Pat. Nos. 6,583,170; 6,479,535; WO 1999/024027; U.S.Pat. No. 6,005,102; EP 0802184; U.S. Pat. Nos. 5,998,402; 5,780,497,5,880,137, WO 2012/048058 and WO 2007/087684.

In another embodiment, the isolated Compound 1 Pattern 1 describedherein can be administered in an effective amount for the treatment ofabnormal tissue of the male reproductive system such as prostate ortesticular cancer, in combination or alternation with an effectiveamount of an androgen (such as testosterone) inhibitor including but notlimited to a selective androgen receptor modulator, a selective androgenreceptor degrader, a complete androgen receptor degrader, or anotherform of partial or complete androgen antagonist. In one embodiment, theprostate or testicular cancer is androgen-resistant. Non-limitingexamples of anti-androgen compounds are provided in WO 2011/156518 andU.S. Pat. Nos. 8,455,534 and 8,299,112. Additional non-limiting examplesof anti-androgen compounds include: enzalutamide, apalutamide,cyproterone acetate, chlormadinone acetate, spironolactone, canrenone,drospirenone, ketoconazole, topilutamide, abiraterone acetate, andcimetidine.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of abirateroneacetate (Zytiga) for the treatment of abnormal tissue of the malereproductive system.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of abirateroneacetate (Zytiga) for the treatment of prostate cancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of enzalutamide forthe treatment of prostate cancer.

In one embodiment, the bioactive agent is an ALK inhibitor. Examples ofALK inhibitors include but are not limited to Crizotinib, Alectinib,ceritinib, TAE684 (NVP-TAE684), GSK1838705A, AZD3463, ASP3026,PF-06463922, entrectinib (RXDX-101), and AP26113. In one embodiment, thebioactive agent is an EGFR inhibitor. Examples of EGFR inhibitorsinclude erlotinib (Tarceva), gefitinib (Iressa), afatinib (Gilotrif),rociletinib (CO-1686), osimertinib (Tagrisso), olmutinib (Olita),naquotinib (ASP8273), nazartinib (EGF816), PF-06747775 (Pfizer),icotinib (BPI-2009), neratinib (HKI-272; PB272); avitinib (AC0010),EAI045, tarloxotinib (TH-4000; PR-610), PF-06459988 (Pfizer),tesevatinib (XL647; EXEL-7647; KD-019), transtinib, WZ-3146, WZ8040,CNX-2006, dacomitinib (PF-00299804; Pfizer), brigatinib (Alunbrig),lorlatinib, and PF-06747775 (PF7775).

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of afatinibdimaleate (Gilotrif) for the treatment of non-small cell lung cancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of alectinib(Alecensa) for the treatment of non-small cell lung cancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of ceritinib(Zykadia) for the treatment of non-small cell lung cancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of crizotinib(Xalkori) for the treatment of non-small cell lung cancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of osimertinib(Tagrisso) for the treatment of non-small cell lung cancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of brigatinib(Alunbrig) for the treatment of non-small cell lung cancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of lorlatinib forthe treatment of non-small cell lung cancer.

In one embodiment, the bioactive agent is an HER-2 inhibitor. Examplesof HER-2 inhibitors include trastuzumab, lapatinib, ado-trastuzumabemtansine, and pertuzumab.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of lapatinibditosylate for the treatment of breast cancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of lapatinibditosylate for the treatment of HER2+ breast cancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of PF7775 for thetreatment of non-small cell lung cancer.

In one embodiment, the bioactive agent is a CD20 inhibitor. Examples ofCD20 inhibitors include obinutuzumab, rituximab, fatumumab, ibritumomab,tositumomab, and ocrelizumab.

In one embodiment, the bioactive agent is a JAK3 inhibitor. Examples ofJAK3 inhibitors include tasocitinib.

In one embodiment, the bioactive agent is a BCL-2 inhibitor. Examples ofBCL-2 inhibitors include venetoclax, ABT-199(4-[4-[[2-(4-Chlorophenyl)-4,4-dimethylcyclohex-1-en-1-yl]methyl]piperazin-1-yl]-N-[[3-nitro-4-[[(tetrahydro-2H-pyran-4-yl)methyl]amino]phenyl]sulfonyl]-2-[(1H-pyrrolo[2,3-b]pyridin-5-yl)oxy]benzamide),ABT-737(4-[4-[[2-(4-chlorophenyl)phenyl]methyl]piperazin-1-yl]-N-[4-[[(2R)-4-(dimethylamino)-1-phenylsulfanylbutan-2-yl]amino]-3-nitrophenyl]sulfonylbenzamide)(navitoclax), ABT-263((R)-4-(4-((4′-chloro-4,4-dimethyl-3,4,5,6-tetrahydro-[1,1′-biphenyl]-2-yl)methyl)piperazin-1-yl)-N-((4-((4-morpholino-1-(phenylthio)butan-2-yl)amino)-3((trifluoromethyl)sulfonyl)phenyl)sulfonyl)benzamide),GX15-070 (obatoclax mesylate,(2Z)-2-[(5Z)-5-[(3,5-dimethyl-1H-pyrrol-2-yl)methylidene]-4-methoxypyrrol-2-ylidene]indole;methanesulfonic acid))), 2-methoxy-antimycin A3, YC137(4-(4,9-dioxo-4,9-dihydronaphtho[2,3-d]thiazol-2-ylamino)-phenyl ester),pogosin, ethyl2-amino-6-bromo-4-(1-cyano-2-ethoxy-2-oxoethyl)-4H-chromene-3-carboxylate,Nilotinib-d3, TW-37(N-[4-[[2-(1,1-Dimethylethyl)phenyl]sulfonyl]phenyl]-2,3,4-trihydroxy-5-[[2-(1-methylethyl)phenyl]methyl]benzamide),Apogossypolone (ApoG2), HA14-1, AT101, sabutoclax, gambogic acid, orG3139 (Oblimersen).

In one aspect, a treatment regimen is provided comprising theadministration of Compound 1 Pattern 1 in combination with at least oneadditional chemotherapeutic agent. The combinations disclosed herein canbe administered for beneficial, additive, or synergistic effect in thetreatment of abnormal cellular proliferative disorders.

In specific embodiments, the treatment regimen includes theadministration of isolated Compound 1 Pattern 1 in combination with atleast one kinase inhibitor. In one embodiment, the at least one kinaseinhibitor is selected from a phosphoinositide 3-kinase (PI3K) inhibitor,a Bruton's tyrosine kinase (BTK) inhibitor, or a spleen tyrosine kinase(Syk) inhibitor, or a combination thereof.

PI3k inhibitors that may be used in the present invention are wellknown. Examples of PI3 kinase inhibitors include but are not limited toWortmannin, demethoxyviridin, perifosine, idelalisib, Pictilisib,Palomid 529, ZSTK474, PWT33597, CUDC-907, and AEZS-136, duvelisib,GS-9820, BKM120, GDC-0032 (Taselisib),(2-[4-[2-(2-Isopropyl-5-methyl-1,2,4-triazol-3-yl)-5,6-dihydroimidazo[1,2-d][1,4]benzoxazepin-9-yl]pyrazol-1-yl]-2-methylpropanamide),MLN-1117 ((2R)-1-Phenoxy-2-butanyl hydrogen (S)-methylphosphonate; orMethyl(oxo) {[(2R)-1-phenoxy-2-butanyl]oxy}phosphonium)), BYL-719((2S)-N1-[4-Methyl-5-[2-(2,2,2-trifluoro-1,1-dimethylethyl)-4-pyridinyl]-2-thiazolyl]-1,2-pyrrolidinedicarboxamide),GSK2126458 (2,4-Difluoro-N-{2-(methyloxy)-5-[4-(4-pyridazinyl)-6-quinolinyl]-3-pyridinyl}benzenesulfonamide)(omipalisib), TGX-221((±)-7-Methyl-2-(morpholin-4-yl)-9-(1-phenylaminoethyl)-pyrido[1,2-a]-pyrimidin-4-one),GSK2636771(2-Methyl-1-(2-methyl-3-(trifluoromethyl)benzyl)-6-morpholino-1H-benzo[d]imidazole-4-carboxylicacid dihydrochloride), KIN-193((R)-2-((1-(7-methyl-2-morpholino-4-oxo-4H-pyrido[1,2-a]pyrimidin-9-yl)ethyl)amino)benzoicacid), TGR-1202/RP5264, GS-9820((S)-1-(4-((2-(2-aminopyrimidin-5-yl)-7-methyl-4-mohydroxypropan-1-one),GS-1101(5-fluoro-3-phenyl-2-([S)]-1-[9H-purin-6-ylamino]-propyl)-3H-quinazolin-4-one),AMG-319, GSK-2269557, SAR245409(N-(4-(N-(3-((3,5-dimethoxyphenyl)amino)quinoxalin-2-yl)sulfamoyl)phenyl)-3-methoxy-4methylbenzamide), BAY80-6946(2-amino-N-(7-methoxy-8-(3-morpholinopropoxy)-2,3-dihydroimidazo[1,2-c]quinaz),AS 252424(5-[1-[5-(4-Fluoro-2-hydroxy-phenyl)-furan-2-yl]-meth-(Z)-ylidene]-thiazolidine-2,4-dione),CZ 24832(5-(2-amino-8-fluoro-[1,2,4]triazolo[1,5-a]pyridin-6-yl)-N-tert-butylpyridine-3-sulfonamide),Buparlisib(5-[2,6-Di(4-morpholinyl)-4-pyrimidinyl]-4-(trifluoromethyl)-2-pyridinamine),GDC-0941(2-(1H-Indazol-4-yl)-6-[[4-(methylsulfonyl)-1-piperazinyl]methyl]-4-(4-morpholinyl)thieno[3,2-d]pyrimidine),GDC-0980((S)-1-(4-((2-(2-aminopyrimidin-5-yl)-7-methyl-4-morpholinothieno[3,2-d]pyrimidin-6yl)methyl)piperazin-1-yl)-2-hydroxypropan-1-one (also known as RG7422)),SF1126((8S,14S,17S)-14-(carboxymethyl)-8-(3-guanidinopropyl)-17-(hydroxymethyl)-3,6,9,12,15-pentaoxo-1-(4-(4-oxo-8-phenyl-4H-chromen-2-yl)morpholino-4-ium)-2-oxa-7,10,13,16-tetraazaoctadecan-18-oate),PF-05212384(N-[4-[[4-(Dimethylamino)-1-piperidinyl]carbonyl]phenyl]-N′-[4-(4,6-di-4-morpholinyl-1,3,5-triazin-2-yl)phenyl]urea)(gedatolisib), LY3023414, BEZ235 (2-Methyl-2-{4-[3methyl-2-oxo-8-(quinolin-3-yl)-2,3-dihydro-1H-imidazo[4,5-c]quinolin-1-yl]phenyl}propanenitrile)(dactolisib), XL-765(N-(3-(N-(3-(3,5-dimethoxyphenylamino)quinoxalin-2-yl)sulfamoyl)phenyl)-3-methoxy-4-methylbenzamide),and GSK1059615(5-[[4-(4-Pyridinyl)-6-quinolinyl]methylene]-2,4-thiazolidenedione),PX886([(3aR,6E,9S,9aR,10R,11aS)-6-[[bis(prop-2-enyl)amino]methylidene]-5-hydroxy-9-(methoxymethyl)-9a,11a-dimethyl-1,4,7-trioxo-2,3,3a,9,10,11-hexahydroindeno[4,5h]isochromen-10-yl]acetate(also known as sonolisib)) LY294002, AZD8186, PF-4989216, pilaralisib,GNE-317, PI-3065, PI-103, NU7441 (KU-57788), HS 173, VS-5584 (SB2343),CZC24832, TG100-115, A66, YM201636, CAY10505, PIK-75, PIK-93, AS-605240,BGT226 (NVP-BGT226), AZD6482, voxtalisib, alpelisib, IC-87114,TGI100713, CH5132799, PKI-402, copanlisib (BAY 80-6946), XL 147, PIK-90,PIK-293, PIK-294, 3-MA (3-methyladenine), AS-252424, AS-604850,apitolisib (GDC-0980; RG7422), and the structures described inWO2014/071109. In one embodiment, isolated Compound 1 Pattern 1 iscombined in a single dosage form with the PIk3 inhibitor.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of alpelisib forthe treatment of solid tumors.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of alpelisib forthe treatment of abnormal tissue of the female reproductive system.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of alpelisib forthe treatment of breast cancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of copanlisibhydrochloride (Aliqopa) for the treatment of lymphoma.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of copanlisibhydrochloride (Aliqopa) for the treatment of follicular lymphoma.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of idelalisib(Zydelig) for the treatment of chronic lymphocytic leukemia.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of idelalisib(Zydelig) for the treatment of Non-Hodgkin lymphoma, includingfollicular B-cell non-Hodgkin lymphoma or small lymphocytic lymphoma.

BTK inhibitors for use in the present invention are well known. Examplesof BTK inhibitors include ibrutinib (also known asPCI-32765)(Imbruvica™)(1-[(3R)-3-[4-amino-3-(4-phenoxy-phenyl)pyrazolo[3,4-d]pyrimidin-1-yl]piperidin-1-yl]prop-2-en-1-one),dianilinopyrimidine-based inhibitors such as AVL-101 and AVL-291/292(N-(3-((5-fluoro-2-((4-(2-methoxyethoxy)phenyl)amino)pyrimidin-4-yl)amino)phenyl)acrylamide) (Avila Therapeutics) (see US Patent Publication No 2011/0117073,incorporated herein in its entirety), Dasatinib([N-(2-chloro-6-methylphenyl)-2-(6-(4-(2-hydroxyethyl)piperazin-1-yl)-2-methylpyrimidin-4-ylamino)thiazole-5-carboxamide],LFM-A13(alpha-cyano-beta-hydroxy-beta-methyl-N-(2,5-ibromophenyl)propenamide),GDC-0834([R-N-(3-(6-(4-(1,4-dimethyl-3-oxopiperazin-2-yl)phenylamino)-4-methyl-5-oxo-4,5-dihydropyrazin-2-yl)-2-methylphenyl)-4,5,6,7-tetrahydrobenzo[b]thiophene-2-carboxamide],CGI-5604-(tert-butyl)-N-(3-(8-(phenylamino)imidazo[1,2-a]pyrazin-6-yl)phenyl)benzamide,CGI-1746(4-(tert-butyl)-N-(2-methyl-3-(4-methyl-6-((4-(morpholine-4-carbonyl)phenyl)amino)-5-oxo-4,5-dihydropyrazin-2-yl)phenyl)benzamide),CNX-774(4-(4-(4-((4-((3-acrylamidophenyl)amino)-5-fluoropyrimidin-2-yl)amino)phenoxy)-N-methylpicolinamide),CTA056(7-benzyl-1-(3-(piperidin-1-yl)propyl)-2-(4-(pyridin-4-yl)phenyl)-1H-imidazo[4,5-g]quinoxalin-6(5H)-one),GDC-0834((R)-N-(3-(6-((4-(1,4-dimethyl-3-oxopiperazin-2-yl)phenyl)amino)-4-methyl-5-oxo-4,5-dihydropyrazin-2-yl)-2-methylphenyl)-4,5,6,7-tetrahydrobenzo[b]thiophene-2-carboxamide),GDC-0837((R)-N-(3-(6-((4-(1,4-dimethyl-3-oxopiperazin-2-yl)phenyl)amino)-4-methyl-5-oxo-4,5-dihydropyrazin-2-yl)-2-methylphenyl)-4,5,6,7-tetrahydrobenzo[b]thiophene-2-carboxamide),HM-71224, ACP-196, ONO-4059 (Ono Pharmaceuticals), PRT062607(4-((3-(2H-1,2,3-triazol-2-yl)phenyl)amino)-2-(((1R,2S)-2-aminocyclohexyl)amino)pyrimidine-5-carboxamidehydrochloride), QL-47(1-(1-acryloylindolin-6-yl)-9-(1-methyl-1H-pyrazol-4-yl)benzo[h][1,6]naphthyridin-2(1H)-one),and RN486(6-cyclopropyl-8-fluoro-2-(2-hydroxymethyl-3-{1-methyl-5-[5-(4-methyl-piperazin-1-yl)-pyridin-2-ylamino]-6-oxo-1,6-dihydro-pyridin-3-yl}-phenyl)-2H-isoquinolin-1-one),and other molecules capable of inhibiting BTK activity, for examplethose BTK inhibitors disclosed in Akinleye et ah, Journal of Hematology& Oncology, 2013, 6:59, the entirety of which is incorporated herein byreference. In one embodiment, an effective amount of the isolatedCompound 1 Pattern 1 is combined in a single dosage form with the BTKinhibitor.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of ibrutinib(Imbruvica) for the treatment of chronic lymphocytic leukemia.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of ibrutinib(Imbruvica) for the treatment of lymphoma, including small lymphocyticlymphoma, mantle cell lymphoma, marginal zone lymphoma, or Waldenströmmacroglobulinemia.

Syk inhibitors for use in the present invention are well known, andinclude, for example, Cerdulatinib(4-(cyclopropylamino)-2-((4-(4-(ethylsulfonyl)piperazin-1-yl)phenyl)amino)pyrimidine-5-carboxamide),entospletinib(6-(1H-indazol-6-yl)-N-(4-morpholinophenyl)imidazo[1,2-a]pyrazin-8-amine),fostamatinib([6-({5-Fluoro-2-[(3,4,5-trimethoxyphenyl)amino]-4-pyrimidinyl}amino)-2,2-dimethyl-3-oxo-2,3-dihydro-4H-pyrido[3,2-b][1,4]oxazin-4-yl]methyldihydrogen phosphate), fostamatinib disodium salt (sodium(6-((5-fluoro-2-((3,4,5-trimethoxyphenyl)amino)pyrimidin-4-yl)amino)-2,2-dimethyl-3-oxo-2H-pyrido[3,2-b][1,4]oxazin-4(3H)-yl)methylphosphate), BAY 61-3606(2-(7-(3,4-Dimethoxyphenyl)-imidazo[1,2-c]pyrimidin-5-ylamino)-nicotinamideHCl), RO9021(6-[(1R,2S)-2-Amino-cyclohexylamino]-4-(5,6-dimethyl-pyridin-2-ylamino)-pyridazine-3-carboxylicacid amide), imatinib (Gleevac;4-[(4-methylpiperazin-1-yl)methyl]-N-(4-methyl-3-{[4-(pyridin-3-yl)pyrimidin-2-yl]amino}phenyl)benzamide),staurosporine, GSK143(2-(((3R,4R)-3-aminotetrahydro-2H-pyran-4-yl)amino)-4-(p-tolylamino)pyrimidine-5-carboxamide),PP2(1-(tert-butyl)-3-(4-chlorophenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine),PRT-060318(2-(((1R,2S)-2-aminocyclohexyl)amino)-4-(m-tolylamino)pyrimidine-5-carboxamide),PRT-062607(4-((3-(2H-1,2,3-triazol-2-yl)phenyl)amino)-2-(((1R,2S)-2-aminocyclohexyl)amino)pyrimidine-5-carboxamidehydrochloride), R112(3,3′-((5-fluoropyrimidine-2,4-diyl)bis(azanediyl))diphenol), R348(3-Ethyl-4-methylpyridine), R406(6-((5-fluoro-2-((3,4,5-trimethoxyphenyl)amino)pyrimidin-4-yl)amino)-2,2-dimethyl-2H-pyrido[3,2-b][1,4]oxazin-3(4H)-one), piceatannol (3-Hydroxyresveratol), YM193306(see Singh et al.Discovery and Development of Spleen Tyrosine Kinase (SYK) Inhibitors, J.Med. Chem. 2012, 55, 3614-3643), 7-azaindole, piceatannol, ER-27319 (seeSingh et al. Discovery and Development of Spleen Tyrosine Kinase (SYK)Inhibitors, J. Med. Chem. 2012, 55, 3614-3643 incorporated in itsentirety herein), Compound D (see Singh et al. Discovery and Developmentof Spleen Tyrosine Kinase (SYK) Inhibitors, J. Med. Chem. 2012, 55,3614-3643 incorporated in its entirety herein), PRT060318 (see Singh etal. Discovery and Development of Spleen Tyrosine Kinase (SYK)Inhibitors, J. Med. Chem. 2012, 55, 3614-3643 incorporated in itsentirety herein), luteolin (see Singh et al. Discovery and Developmentof Spleen Tyrosine Kinase (SYK) Inhibitors, J. Med. Chem. 2012, 55,3614-3643 incorporated in its entirety herein), apigenin (see Singh etal. Discovery and Development of Spleen Tyrosine Kinase (SYK)Inhibitors, J. Med. Chem. 2012, 55, 3614-3643 incorporated in itsentirety herein), quercetin (see Singh et al. Discovery and Developmentof Spleen Tyrosine Kinase (SYK) Inhibitors, J. Med. Chem. 2012, 55,3614-3643 incorporated in its entirety herein), fisetin (see Singh etal. Discovery and Development of Spleen Tyrosine Kinase (SYK)Inhibitors, J. Med. Chem. 2012, 55, 3614-3643 incorporated in itsentirety herein), myricetin (see Singh et al. Discovery and Developmentof Spleen Tyrosine Kinase (SYK) Inhibitors, J. Med. Chem. 2012, 55,3614-3643 incorporated in its entirety herein), morin (see Singh et al.Discovery and Development of Spleen Tyrosine Kinase (SYK) Inhibitors, J.Med. Chem. 2012, 55, 3614-3643 incorporated in its entirety herein). Inone embodiment an effective amount of the isolated Compound 1 Pattern 1is combined in a single dosage form with the Syk inhibitor.

In one embodiment, the at least one additional chemotherapeutic agent isa protein cell death-1 (PD-1) inhibitor. PD-1 inhibitors are known inthe art, and include, for example, nivolumab (BMS), pembrolizumab(Merck), pidilizumab (CureTech/Teva), AMP-244 (Amplimmune/GSK),BMS-936559 (BMS), and MEDI4736 (Roche/Genentech). In one embodiment, aneffective amount of the isolated Compound 1 Pattern 1 is combined in asingle dosage form with the PD-1 inhibitor.

In one embodiment, the at least one additional chemotherapeutic agent isa B-cell lymphoma 2 (Bcl-2) protein inhibitor. BCL-2 inhibitors areknown in the art, and include, for example, ABT-199(4-[4-[[2-(4-Chlorophenyl)-4,4-dimethylcyclohex-1-en-1-yl]methyl]piperazin-1-yl]-N-[[3-nitro-4-[[(tetrahydro-2H-pyran-4-yl)methyl]amino]phenyl]sulfonyl]-2-[(1H-pyrrolo[2,3-b]pyridin-5-yl)oxy]benzamide),ABT-737(4-[4-[[2-(4-chlorophenyl)phenyl]methyl]piperazin-1-yl]-N-[4-[[(2R)-4-(dimethylamino)-1-phenylsulfanylbutan-2-yl]amino]-3-nitrophenyl]sulfonylbenzamide),ABT-263((R)-4-(4-((4′-chloro-4,4-dimethyl-3,4,5,6-tetrahydro-[1,1′-biphenyl]-2-yl)methyl)piperazin-1-yl)-N-((4-((4-morpholino-1-(phenylthio)butan-2-yl)amino)-3((trifluoromethyl)sulfonyl)phenyl)sulfonyl)benzamide),GX15-070 (obatoclax mesylate,(2Z)-2-[(5Z)-5-[(3,5-dimethyl-1H-pyrrol-2-yl)methylidene]-4-methoxypyrrol-2-ylidene]indole;methanesulfonic acid))), 2-methoxy-antimycin A3, YC137(4-(4,9-dioxo-4,9-dihydronaphtho[2,3-d]thiazol-2-ylamino)-phenyl ester),pogosin, ethyl2-amino-6-bromo-4-(1-cyano-2-ethoxy-2-oxoethyl)-4H-chromene-3-carboxylate,Nilotinib-d3, TW-37(N-[4-[[2-(1,1-Dimethylethyl)phenyl]sulfonyl]phenyl]-2,3,4-trihydroxy-5-[[2-(1-methylethyl)phenyl]methyl]benzamide),Apogossypolone (ApoG2), or G3139 (Oblimersen). In one embodiment, aneffective amount of the isolated Compound 1 Pattern 1 is combined in asingle dosage form with the at least one BCL-2 inhibitor.

In one embodiment, a combination described herein can be furthercombined with an additional therapeutic to treat the cancer. The secondtherapy can be an immunotherapy. As discussed in more detail below, aneffective amount of the isolated Compound 1 Pattern 1 can be conjugatedto an antibody, radioactive agent, or other targeting agent that directsthe compound to the diseased or abnormally proliferating cell. Inanother embodiment, the combination is used in combination with anotherpharmaceutical or a biologic agent (for example an antibody) to increasethe efficacy of treatment with a combined or a synergistic approach. Inan embodiment, combination can be used with T-cell vaccination, whichtypically involves immunization with inactivated autoreactive T cells toeliminate a cancer cell population as described herein. In anotherembodiment, the combination is used in combination with a bispecificT-cell Engager (BiTE), which is an antibody designed to simultaneouslybind to specific antigens on endogenous T cells and cancer cells asdescribed herein, linking the two types of cells.

In one embodiment, the bioactive agent is a MEK inhibitor. MEKinhibitors are well known, and include, for example,trametinib/GSK1120212(N-(3-{3-Cyclopropyl-5-[(2-fluoro-4-iodophenyl)amino]-6,8-dimethyl-2,4,7-trioxo-3,4,6,7-tetrahydropyrido[4,3-d]pyrimidin-1(2H-yl}phenyl)acetamide),selumetinib(6-(4-bromo-2-chloroanilino)-7-fluoro-N-(2-hydroxyethoxy)-3-methylbenzimidazole-5-carboxamide),pimasertib/AS703026/MSC 1935369((S)-N-(2,3-dihydroxypropyl)-3-((2-fluoro-4-iodophenyl)amino)isonicotinamide),XL-518/GDC-0973(1-({3,4-difluoro-2-[(2-fluoro-4-iodophenyl)amino]phenyl}carbonyl)-3-[(2S)-piperidin-2-yl]azetidin-3-ol),refametinib/BAY869766/RDEA1 19(N-(3,4-difluoro-2-(2-fluoro-4-iodophenylamino)-6-methoxyphenyl)-1-(2,3-dihydroxypropyl)cyclopropane-1-sulfonamide),PD-0325901 (N-[(2R)-2,3-Dihydroxypropoxy]-3,4-difluoro-2-[(2-fluoro-4-iodophenyl)amino]-benzamide), TAK733((R)-3-(2,3-Dihydroxypropyl)-6-fluoro-5-(2-fluoro-4-iodophenylamino)-8-methylpyrido[2,3-d]pyrimidine-4,7(3H,8H)-dione),MEK162/ARRY438162(5-[(4-Bromo-2-fluorophenyl)amino]-4-fluoro-N-(2-hydroxyethoxy)-1-methyl-1H-benzimidazole-6-carboxamide),R05126766(3-[[3-Fluoro-2-(methylsulfamoylamino)-4-pyridyl]methyl]-4-methyl-7-pyrimidin-2-yloxychromen-2-one),WX-554, R04987655/CH4987655(3,4-difluoro-2-((2-fluoro-4-iodophenyl)amino)-N-(2-hydroxyethoxy)-5-((3-oxo-1,2-oxazinan-2yl)methyl)benzamide),or AZD8330 (2-((2-fluoro-4-iodophenyl)amino)-N-(2hydroxyethoxy)-1,5-dimethyl-6-oxo-1,6-dihydropyridine-3-carboxamide),U0126-EtOH, PD184352 (CI-1040), GDC-0623, BI-847325, cobimetinib,PD98059, BIX 02189, BIX 02188, binimetinib, SL-327, TAK-733, PD318088.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of binimetinib forthe treatment of melanoma, including BRAF-mutant melanoma andNRAS-mutant melanoma.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of cobimetinib(Cotellic) for the treatment of melanoma, including BRAF-mutant melanomaand NRAS-mutant melanoma.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of binimetinib forthe treatment of ovarian cancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of selumetinib forthe treatment of non-small cell lung cancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of selumetinib forthe treatment of thyroid cancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of trametinib(Mekinist) for the treatment of thyroid cancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of trametinib(Mekinist) for the treatment of melanoma.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of trametinib(Mekinist) for the treatment of non-small cell lung cancer.

In one embodiment, the bioactive agent is a Raf inhibitor. Rafinhibitors are known and include, for example, Vemurafinib(N-[3-[[5-(4-Chlorophenyl)-1H-pyrrolo[2,3-b]pyridin-3-yl]carbonyl]-2,4-difluorophenyl]-1-propanesulfonamide),sorafenib tosylate(4-[4-[[4-chloro-3-(trifluoromethyl)phenyl]carbamoylamino]phenoxy]-N-methylpyridine-2-carboxamide;4-methylbenzenesulfonate),AZ628(3-(2-cyanopropan-2-yl)-N-(4-methyl-3-(3-methyl-4-oxo-3,4-dihydroquinazolin-6-ylamino)phenyl)benzamide),NVP-BHG712(4-methyl-3-(1-methyl-6-(pyridin-3-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-ylamino)-N-(3-(trifluoromethyl)phenyl)benzamide),RAF-265(1-methyl-5-[2-[5-(trifluoromethyl)-1H-imidazol-2-yl]pyridin-4-yl]oxy-N-[4-(trifluoromethyl)phenyl]benzimidazol-2-amine),2-Bromoaldisine(2-Bromo-6,7-dihydro-1H,5H-pyrrolo[2,3-c]azepine-4,8-dione), Raf KinaseInhibitor IV(2-chloro-5-(2-phenyl-5-(pyridin-4-yl)-1H-imidazol-4-yl)phenol),Sorafenib N-Oxide(4-[4-[[[[4-Chloro-3(trifluoroMethyl)phenyl]aMino]carbonyl]aMino]phenoxy]-N-Methyl-2pyridinecarboxaMide1-Oxide), PLX-4720, dabrafenib (GSK2118436), GDC-0879, RAF265, AZ 628,SB590885, ZM336372, GW5074, TAK-632, CEP-32496, LY3009120, and GX818(Encorafenib).

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of dabrafenib(Tafinlar) for the treatment of thyroid cancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of dabrafenib(Tafinlar) for the treatment of melanoma.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of dabrafenib(Tafinlar) for the treatment of non-small cell lung cancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of encorafenib forthe treatment of melanoma, including BRAF-mutant melanoma.

In one embodiment, the additional therapy is a monoclonal antibody(MAb). Some MAbs stimulate an immune response that destroys cancercells. Similar to the antibodies produced naturally by B cells, theseMAbs “coat” the cancer cell surface, triggering its destruction by theimmune system. For example, bevacizumab targets vascular endothelialgrowth factor(VEGF), a protein secreted by tumor cells and other cellsin the tumor's microenvironment that promotes the development of tumorblood vessels. When bound to bevacizumab, VEGF cannot interact with itscellular receptor, preventing the signaling that leads to the growth ofnew blood vessels. Similarly, cetuximab and panitumumab target theepidermal growth factor receptor (EGFR), and trastuzumab targets thehuman epidermal growth factor receptor 2 (HER-2). MAbs that bind to cellsurface growth factor receptors prevent the targeted receptors fromsending their normal growth-promoting signals. They may also triggerapoptosis and activate the immune system to destroy tumor cells.

Another group of cancer therapeutic MAbs are the immunoconjugates. TheseMAbs, which are sometimes called immunotoxins or antibody-drugconjugates, consist of an antibody attached to a cell-killing substance,such as a plant or bacterial toxin, a chemotherapy drug, or aradioactive molecule. The antibody latches onto its specific antigen onthe surface of a cancer cell, and the cell-killing substance is taken upby the cell. FDA-approved conjugated MAbs that work this way includeado-trastuzumab emtansine, which targets the HER-2 molecule to deliverthe drug DM1, which inhibits cell proliferation, to HER-2 expressingmetastatic breast cancer cells.

Immunotherapies with T cells engineered to recognize cancer cells viabispecific antibodies (bsAbs) or chimeric antigen receptors (CARs) areapproaches with potential to ablate both dividing and non/slow-dividingsubpopulations of cancer cells.

Bispecific antibodies, by simultaneously recognizing target antigen andan activating receptor on the surface of an immune effector cell, offeran opportunity to redirect immune effector cells to kill cancer cells.The other approach is the generation of chimeric antigen receptors byfusing extracellular antibodies to intracellular signaling domains.Chimeric antigen receptor-engineered T cells are able to specificallykill tumor cells in a MHC-independent way.

In some embodiments, the combination can be administered to the subjectin further combination with other chemotherapeutic agents. Ifconvenient, the combination described herein can be administered at thesame time as another chemotherapeutic agent, in order to simplify thetreatment regimen. In some embodiments, the combination and the otherchemotherapeutic can be provided in a single formulation. In oneembodiment, the use of the compounds described herein is combined in atherapeutic regime with other agents. Such agents may include, but arenot limited to, tamoxifen, midazolam, letrozole, bortezomib,anastrozole, goserelin, an mTOR inhibitor, a PI3 kinase inhibitors, dualmTOR-PI3K inhibitors, MEK inhibitors, RAS inhibitors, ALK inhibitors,HSP inhibitors (for example, HSP70 and HSP 90 inhibitors, or acombination thereof), BCL-2 inhibitors, apopototic inducing compounds,AKT inhibitors, including but not limited to, MK-2206, GSK690693,Perifosine, (KRX-0401), GDC-0068, Triciribine, AZD5363, Honokiol,PF-04691502, and ipatasertib, Miltefosine; PD-1 inhibitors including butnot limited to, Nivolumab, CT-011, MK-3475, BMS936558, and AMP-514 orFLT-3 inhibitors, including but not limited to, P406, Dovitinib,Quizartinib (AC220), Amuvatinib (MP-470), Tandutinib (MLN518),ENMD-2076, and KW-2449, or combinations thereof.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of ipatasertib forthe treatment of breast cancer, including triple negative breast cancer.

In one embodiment, the bioactive agent is an mTOR inhibitor. Examples ofmTOR inhibitors include but are not limited to vistusertib and rapamycinand its analogs, everolimus (Afinitor), temsirolimus, ridaforolimus,sirolimus, and deforolimus. Examples of MEK inhibitors include but arenot limited to tametinib/GSK1120212(N-(3-{3-Cyclopropyl-5-[(2-fluoro-4-iodophenyl)amino]-6,8-dimethyl-2,4,7-trioxo-3,4,6,7-tetrahydropyrido[4,3-d]pyrimidin-1(2H-yl}phenyl)acetamide),selumetinob(6-(4-bromo-2-chloroanilino)-7-fluoro-N-(2-hydroxyethoxy)-3-methylbenzimidazole-5-carboxamide),pimasertib/AS703026/MSC1935369((S)-N-(2,3-dihydroxypropyl)-3-((2-fluoro-4-iodophenyl)amino)isonicotinamide),XL-518/GDC-0973(1-({3,4-difluoro-2-[(2-fluoro-4-iodophenyl)amino]phenyl}carbonyl)-3-[(2S)-piperidin-2-yl]azetidin-3-ol),refametinib/BAY869766/RDEA119(N-(3,4-difluoro-2-(2-fluoro-4-iodophenylamino)-6-methoxyphenyl)-1-(2,3-dihydroxypropyl)cyclopropane-1-sulfonamide),PD-0325901(N-[(2R)-2,3-Dihydroxypropoxy]-3,4-difluoro-2-[(2-fluoro-4-iodophenyl)amino]-benzamide),TAK733((R)-3-(2,3-Dihydroxypropyl)-6-fluoro-5-(2-fluoro-4-iodophenylamino)-8-methylpyrido[2,3d]pyrimidine-4,7(3H,8H)-dione),MEK162/ARRY438162(5-[(4-Bromo-2-fluorophenyl)amino]-4-fluoro-N-(2-hydroxyethoxy)-1-methyl-1H-benzimidazole-6carboxami de), R05126766(3-[[3-Fluoro-2-(methylsulfamoylamino)-4-pyridyl]methyl]-4-methyl-7-pyrimidin-2-yloxychromen-2-one),WX-554, R04987655/CH4987655(3,4-difluoro-2-((2-fluoro-4-iodophenyl)amino)-N-(2-hydroxyethoxy)-5-((3-oxo-1,2-oxazinan-2yl)methyl)benzamide), or AZD8330(2-((2-fluoro-4-iodophenyl)amino)-N-(2-hydroxyethoxy)-1,5-dimethyl-6-oxo-1,6-dihydropyridine-3-carboxamide).

In one embodiment, the bioactive agent is a RAS inhibitor. Examples ofRAS inhibitors include but are not limited to Reolysin and siG12D LODER.

In one embodiment, the bioactive agent is an ALK inhibitor. Examples ofALK inhibitors include but are not limited to Crizotinib, AP26113, andLDK378.

In one embodiment, the bioactive agent is a HSP inhibitor. HSPinhibitors include but are not limited to Geldanamycin or17-N-Allylamino-17-demethoxygeldanamycin (17AAG), and Radicicol. In aparticular embodiment, a compound described herein is administered incombination with letrozole and/or tamoxifen. Other chemotherapeuticagents that can be used in combination with the compounds describedherein include, but are not limited to, chemotherapeutic agents that donot require cell cycle activity for their anti-neoplastic effect.

Additional bioactive compounds include, for example, everolimus,trabectedin, abraxane, TLK 286, AV-299, DN-101, pazopanib, GSK690693,RTA 744, ON 0910.Na, AZD 6244 (ARRY-142886), AMN-107, TKI-258,GSK461364, AZD 1152, enzastaurin, vandetanib, ARQ-197, MK-0457, MLN8054,PHA-739358, R-763, AT-9263, a FLT-3 inhibitor, a VEGFR inhibitor, anaurora kinase inhibitor, a PIK-1 modulator, an HDAC inhibitor, a c-METinhibitor, a PARP inhibitor, a Cdk inhibitor, an IGFR-TK inhibitor, ananti-HGF antibody, a focal adhesion kinase inhibitor, a Map kinase (mek)inhibitor, a VEGF trap antibody, pemetrexed, panitumumab, amrubicin,oregovomab, Lep-etu, nolatrexed, azd2171, batabulin, ofatumumab,zanolimumab, edotecarin, tetrandrine, rubitecan, tesmilifene,oblimersen, ticilimumab, ipilimumab, gossypol, Bio 111, 131-I-TM-601,ALT-110, BIO 140, CC 8490, cilengitide, gimatecan, IL13-PE38QQR, INO1001, IPdR₁ KRX-0402, lucanthone, LY317615, neuradiab, vitespan, Rta744, Sdx 102, talampanel, atrasentan, Xr 311, romidepsin, ADS-100380,sunitinib, 5-fluorouracil, vorinostat, etoposide, gemcitabine,doxorubicin, liposomal doxorubicin, 5′-deoxy-5-fluorouridine,vincristine, temozolomide, ZK-304709, seliciclib; PD0325901, AZD-6244,capecitabine, L-Glutamic acid,N-[4-[2-(2-amino-4,7-dihydro-4-oxo-1H-pyrrolo[2,3-d]pyrimidin-5-yl)ethyl]benzoyl]-,disodium salt, heptahydrate, camptothecin, PEG-labeled irinotecan,tamoxifen, toremifene citrate, anastrazole, exemestane, letrozole,DES(diethylstilbestrol), estradiol, estrogen, conjugated estrogen,bevacizumab, IMC-1C11, CHIR-258); 3-[5-(methylsulfonylpiperadinemethyl)-indolyl-quinolone, vatalanib, AG-013736,AVE-0005, goserelin acetate, leuprolide acetate, triptorelin pamoate,medroxyprogesterone acetate, hydroxyprogesterone caproate, megestrolacetate, raloxifene, bicalutamide, flutamide, nilutamide, megestrolacetate, CP-724714; TAK-165, HKI-272, erlotinib, lapatanib, canertinib,ABX-EGF antibody, erbitux, EKB-569, PKI-166, GW-572016, Ionafarnib,BMS-214662, tipifarnib; amifostine, NVP-LAQ824, suberoyl analidehydroxamic acid, valproic acid, trichostatin A, FK-228, SU11248,sorafenib, KRN951, aminoglutethimide, amsacrine, anagrelide,L-asparaginase, Bacillus Calmette-Guerin (BCG) vaccine, adriamycin,bleomycin, buserelin, busulfan, carboplatin, carmustine, chlorambucil,cisplatin, cladribine, clodronate, cyproterone, cytarabine, dacarbazine,dactinomycin, daunorubicin, diethylstilbestrol, epirubicin, fludarabine,fludrocortisone, fluoxymesterone, flutamide, gleevec, gemcitabine,hydroxyurea, idarubicin, ifosfamide, imatinib, leuprolide, levamisole,lomustine, mechlorethamine, melphalan, 6-mercaptopurine, mesna,methotrexate, mitomycin, mitotane, mitoxantrone, nilutamide, octreotide,oxaliplatin, pamidronate, pentostatin, plicamycin, porfimer,procarbazine, raltitrexed, rituximab, streptozocin, teniposide,testosterone, thalidomide, thioguanine, thiotepa, tretinoin, vindesine,13-cis-retinoic acid, phenylalanine mustard, uracil mustard,estramustine, altretamine, floxuridine, 5-deooxyuridine, cytosinearabinoside, 6-mecaptopurine, deoxycoformycin, calcitriol, valrubicin,mithramycin, vinblastine, vinorelbine, topotecan, razoxin, marimastat,COL-3, neovastat, BMS-275291, squalamine, endostatin, SU5416, SU6668,EMD121974, interleukin-12, IM862, angiostatin, vitaxin, droloxifene,idoxyfene, spironolactone, finasteride, cimitidine, trastuzumab,denileukin diftitox, gefitinib, bortezimib, paclitaxel, cremophor-freepaclitaxel, docetaxel, epithilone B, BMS-247550, BMS-310705,droloxifene, 4-hydroxytamoxifen, pipendoxifene, ERA-923, arzoxifene,fulvestrant, acolbifene, lasofoxifene, idoxifene, TSE-424, HMR-3339,ZK186619, topotecan, PTK787/ZK 222584, VX-745, PD 184352, rapamycin,40-O-(2-hydroxyethyl)-rapamycin, temsirolimus, AP-23573, RAD001,ABT-578, BC-210, LY294002, LY292223, LY292696, LY293684, LY293646,wortmannin, ZM336372, L-779,450, PEG-filgrastim, darbepoetin,erythropoietin, granulocyte colony-stimulating factor, zolendronate,prednisone, cetuximab, granulocyte macrophage colony-stimulating factor,histrelin, pegylated interferon alfa-2a, interferon alfa-2a, pegylatedinterferon alfa-2b, interferon alfa-2b, azacitidine, PEG-L-asparaginase,lenalidomide, gemtuzumab, hydrocortisone, interleukin-11, dexrazoxane,alemtuzumab, all-transretinoic acid, ketoconazole, interleukin-2,megestrol, immune globulin, nitrogen mustard, methylprednisolone,ibritgumomab tiuxetan, androgens, decitabine, hexamethylmelamine,bexarotene, tositumomab, arsenic trioxide, cortisone, editronate,mitotane, cyclosporine, liposomal daunorubicin, Edwina-asparaginase,strontium 89, casopitant, netupitant, an NK-1 receptor antagonist,palonosetron, aprepitant, diphenhydramine, hydroxyzine, metoclopramide,lorazepam, alprazolam, haloperidol, droperidol, dronabinol,dexamethasone, methylprednisolone, prochlorperazine, granisetron,ondansetron, dolasetron, tropisetron, pegfilgrastim, erythropoietin, aplatelet-derived growth factor receptor alpha (PDGFR-α) antibody,epoetin alfa, darbepoetin alfa and mixtures thereof.

In one embodiment, an effective amount of the isolated Compound 1Pattern 1 described herein can be combined with a PARP inhibitorselected from niraparib tosylate monohydrate (Zejula), olaparib(Lynparza), rucaparib camsylate (Rubraca), and talazoparib.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of niraparibtosylate monohydrate (Zejula) for the treatment of abnormal tissue ofthe female reproductive system, including ovarian epithelial cancer orfallopian tube cancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of niraparibtosylate monohydrate (Zejula) for the treatment of peritoneal cancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of olaparib(Lynparza) for the treatment of abnormal tissue of the femalereproductive system, including breast cancer, ovarian cancer, ovarianepithelial cancer or fallopian tube cancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of olaparib(Lynparza) for the treatment of BRAC1 or BRAC2-mutated breast cancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of olaparib(Lynparza) for the treatment of HER2-breast cancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of olaparib(Lynparza) for the treatment of peritoneal cancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of rucaparibcamsylate (Rubraca) for the treatment of abnormal tissue of the femalereproductive system, including breast cancer, ovarian cancer, ovarianepithelial cancer or fallopian tube cancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of rucaparibcamsylate (Rubraca) for the treatment of peritoneal cancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of talazoparib forthe treatment of abnormal tissue of the female reproductive system,including breast cancer, ovarian cancer, ovarian epithelial cancer orfallopian tube cancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of talazoparib forthe treatment of BRAC1 or BRAC2-mutated breast cancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of olaratumab forthe treatment of soft tissue sarcoma.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of savolitinib forthe treatment of adenocarcinoma.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of savolitinib forthe treatment of non-small cell lung cancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of savolitinib forthe treatment of renal cell carcinoma.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of vistusertib forthe treatment of advanced breast cancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of vistusertib forthe treatment of advanced breast cancer.

In one embodiment, an effective amount of the isolated Compound 1Pattern 1 described herein can be combined with a chemotherapeuticselected from, but are not limited to, Imatinib mesylate (Gleevac®),Dasatinib (Sprycel®), Nilotinib (Tasigna®), Bosutinib (Bosulif®),Trastuzumab (Herceptin®), Pertuzumab (Perjeta™), Lapatinib (Tykerb®),Gefitinib (Iressa®), Erlotinib (Tarceva®), Cetuximab (Erbitux®),Panitumumab (Vectibix®), Vandetanib (Caprelsa®), Vemurafenib(Zelboraf®), Vorinostat (Zolinza®), Romidepsin (Istodax®), Bexarotene(Tagretin®), Alitretinoin (Panretin®), Tretinoin (Vesanoid®),Carfilizomib (Kyprolis™), Pralatrexate (Folotyn®), Bevacizumab(Avastin®), Ziv-aflibercept (Zaltrap®), Sorafenib (Nexavar®), Sunitinib(Sutent®), Pazopanib (Votrient®), Regorafenib (Stivarga®), andCabozantinib (Cometriq™).

In one embodiment, an effective amount of the isolated Compound 1Pattern 1 described herein can be combined with a CD4/6 inhibitorincluding abemaciclib (Versenio), palbociclib (Ibrance), or trilaciclib.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of abemaciclib(Versenio) for the treatment of breast cancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of abemaciclib(Versenio) for the treatment of HR+HER2-breast cancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of palbociclib(Ibrance) for the treatment of breast cancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of palbociclib(Ibrance) for the treatment of HR+HER2-breast cancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of palbociclib(Ibrance) for the treatment of breast cancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of palbociclib(Ibrance) for the treatment of metastatic triple negative breast cancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of palbociclib(Ibrance) for the treatment of small cell lung cancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of cabozantinibS-malate (Cometrig™) for the treatment of thyroid cancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of cabozantinibS-maleate (Cometrig™) for the treatment of renal cell carcinoma.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of dasatinib(Sprycel) for the treatment of leukemia, including acute lymphoblasticleukemia or chronic myelogenous leukemia.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of dasatinib(Sprycel) for the treatment of prostate cancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of Erlotinib(Tarceva®) for the treatment of prostate cancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of Gefitinib(Iressa®) for the treatment of prostate cancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of imatinibmesylate (Gleevec) for the treatment of leukemia, including acutelymphoblastic leukemia, chronic eosinophilic leukemia, hypereosinophilicsyndrome, or chronic myelogenous leukemia.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of trastuzumab(Herceptin) for the treatment of adenocarcinoma.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of trastuzumab(Herceptin) for the treatment of breast cancer, including HER2+ breastcancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of imatinibmesylate (Gleevec) for the treatment of tumors, including but notlimited to dermatofibrosarcoma protuberans and gastrointestinal stromaltumors.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of imatinibmesylate (Gleevec) for the treatment ofmyelodysplastic/myeloproliferative neoplasms.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of imatinibmesylate (Gleevec) for the treatment of systemic mastocytosis.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of nilotinib(Tasigna) for the treatment of chronic myelogenous leukemia, includingPhiladelphia chromosome positive chronic myeloid leukemia (Ph+CML).

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of pazopanibhydrochloride (Votrient) for the treatment of renal cell carcinoma.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of pazopanibhydrochloride (Votrient) for the treatment of soft tissue sarcoma.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of regorafenib(Stivarga) for the treatment of colorectal cancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of regorafenib(Stivarga) for the treatment of gastrointestinal stromal tumor.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of regorafenib(Stivarga) for the treatment of hepatocellular carcinoma.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of sorafenibTosylate (Nexavar) for the treatment of carcinoma, includinghepatocellular carcinoma or renal cell carcinoma.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of sunitinib malate(Sutent) for the treatment of gastrointestinal stromal tumor.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of sunitinib malate(Sutent) for the treatment of pancreatic cancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of sunitinib malate(Sutent) for the treatment of renal cell carcinoma.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of vemurafenib(Zelboraf) for the treatment of Erdheim-Chester disease.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of vemurafenib(Zelboraf) for the treatment of melanoma.

In certain aspects, the additional therapeutic agent is ananti-inflammatory agent, a chemotherapeutic agent, a radiotherapeutic,additional therapeutic agents, or immunosuppressive agents.

Suitable chemotherapeutic agents include, but are not limited to,radioactive molecules, toxins, also referred to as cytotoxins orcytotoxic agents, which includes any agent that is detrimental to theviability of cells, agents, and liposomes or other vesicles containingchemotherapeutic compounds. General anticancer pharmaceutical agentsinclude: Vincristine (Oncovin®) or liposomal vincristine (Marqibo®),Daunorubicin (daunomycin or Cerubidine®) or doxorubicin (Adriamycin®),Cytarabine (cytosine arabinoside, ara-C, or Cytosar®), L-asparaginase(Elspar®) or PEG-L-asparaginase (pegaspargase or Oncaspar®), Etoposide(VP-16), Teniposide (Vumon®), 6-mercaptopurine (6-MP or Purinethol®),Methotrexate, Cyclophosphamide (Cytoxan®), Prednisone, Dexamethasone(Decadron), imatinib (Gleevec®), dasatinib (Sprycel®), nilotinib(Tasigna®), bosutinib (Bosulif®), and ponatinib (Iclusig™) Examples ofadditional suitable chemotherapeutic agents include but are not limitedto 1-dehydrotestosterone, 5-fluorouracil decarbazine, 6-mercaptopurine,6-thioguanine, actinomycin D, adriamycin, aldesleukin, alkylatingagents, allopurinol sodium, altretamine, amifostine, anastrozole,anthramycin (AMC)), anti-mitotic agents, cis-dichlorodiamine platinum(II) (DDP) cisplatin), diamino dichloro platinum, anthracycline, anantibiotic, an antimetabolite, asparaginase, BCG live (intravesical),betamethasone sodium phosphate and betamethasone acetate, bicalutamide,bleomycin sulfate, busulfan, calcium leucouorin, calicheamicin,capecitabine, carboplatin, lomustine (CCNU), carmustine (BSNU),Chlorambucil, Cisplatin, Cladribine, Colchicin, conjugated estrogens,Cyclophosphamide, Cyclothosphamide, Cytarabine, Cytarabine, cytochalasinB, Cytoxan, Dacarbazine, Dactinomycin, dactinomycin (formerlyactinomycin), daunirubicin HCl, daunorucbicin citrate, denileukindiftitox, Dexrazoxane, Dibromomannitol, dihydroxy anthracin dione,Docetaxel, dolasetron mesylate, doxorubicin HCl, dronabinol, E. coliL-asparaginase, emetine, epoetin-α, Erwinia L-asparaginase, esterifiedestrogens, estradiol, estramustine phosphate sodium, ethidium bromide,ethinyl estradiol, etidronate, etoposide citrororum factor, etoposidephosphate, filgrastim, floxuridine, fluconazole, fludarabine phosphate,fluorouracil, flutamide, folinic acid, gemcitabine HCl, glucocorticoids,goserelin acetate, gramicidin D, granisetron HCl, hydroxyurea,idarubicin HCl, ifosfamide, interferon α-2b, irinotecan HCl, letrozole,leucovorin calcium, leuprolide acetate, levamisole HCl, lidocaine,lomustine, maytansinoid, mechlorethamine HCl, medroxyprogesteroneacetate, megestrol acetate, melphalan HCl, mercaptipurine, mesna,methotrexate, methyltestosterone, mithramycin, mitomycin C, mitotane,mitoxantrone, nilutamide, octreotide acetate, ondansetron HCL,paclitaxel, pamidronate disodium, pentostatin, pilocarpine HCl,plimycin, polifeprosan 20 with carmustine implant, porfimer sodium,procaine, procarbazine HCl, propranolol, rituximab, sargramostim,streptozotocin, tamoxifen, taxol, teniposide, tenoposide, testolactone,tetracaine, thioepa chlorambucil, thioguanine, thiotepa, topotecan HCl,toremifene citrate, trastuzumab, tretinoin, valrubicin, vinblastinesulfate, vincristine sulfate, and vinorelbine tartrate.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of bosutinib(Bosulif®) for the treatment of chronic myelogenous leukemia (CML).

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of ponatinibhydrochloride (Iclusig) for the treatment of leukemia, including acutelymphoblastic leukemia and chronic myelogenous leukemia.

Additional therapeutic agents that can be administered in combinationwith a compound disclosed herein can include bevacizumab, sutinib,sorafenib, 2-methoxyestradiol or 2ME2, finasunate, vatalanib,vandetanib, aflibercept, volociximab, etaracizumab (MEDI-522),cilengitide, erlotinib, cetuximab, panitumumab, gefitinib, trastuzumab,dovitinib, figitumumab, atacicept, rituximab, alemtuzumab, aldesleukine,atlizumab, tocilizumab, temsirolimus, everolimus, lucatumumab,dacetuzumab, HLL1, huN901-DM1, atiprimod, natalizumab, bortezomib,carfilzomib, marizomib, tanespimycin, saquinavir mesylate, ritonavir,nelfinavir mesylate, indinavir sulfate, belinostat, panobinostat,mapatumumab, lexatumumab, dulanermin, ABT-737, oblimersen, plitidepsin,talmapimod, P276-00, enzastaurin, tipifarnib, perifosine, imatinib,dasatinib, lenalidomide, thalidomide, simvastatin, celecoxib,bazedoxifene, AZD4547, rilotumumab, oxaliplatin (Eloxatin), PD0332991,ribociclib (LEE011), amebaciclib (LY2835219), HDM201, fulvestrant(Faslodex), exemestane (Aromasin), PIM447, ruxolitinib (INC424), BGJ398,necitumumab, pemetrexed (Alimta), and ramucirumab (IMC-1121B).

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of everolimus(Afinitor) for the treatment of breast cancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of everolimus(Afinitor) for the treatment of HR+, HER2-breast cancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of everolimus(Afinitor) for the treatment of pancreatic cancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of everolimus(Afinitor) for the treatment of gastrointestinal cancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of everolimus(Afinitor) for the treatment of lung cancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of everolimus(Afinitor) for the treatment of renal cell carcinoma.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of everolimus(Afinitor) for the treatment of astrocytoma, including subependymalgiant cell astrocytoma.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of fulvestrant(Faslodex) for the treatment of breast cancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of fulvestrant(Faslodex) for the treatment of HR+, HER2-breast cancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of ramucirumab forthe treatment of adenocarcinoma.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of ramucirumab forthe treatment of non-small cell lung cancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of ramucirumab forthe treatment of colorectal cancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of ribociclib(Kisqali) for the treatment of breast cancer.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of ribociclib(Kisqali) for the treatment of HR+ and HER2-breast cancer.

In one aspect of the present invention, a compound described herein canbe combined with at least one IDH1 or IDH2 inhibitor. In one embodiment,an effective amount of Compound 1 Pattern 1 is administered incombination with an effective amount of enasidenib mesylate (Idhifa) forthe treatment of acute myeloid leukemia.

In one aspect of the present invention, a compound described herein canbe combined with at least one fibroblast growth factor receptor (FGFR)tyrosine kinase inhibitor. In one embodiment, an effective amount ofCompound 1 Pattern 1 is administered in combination with an effectiveamount of erdafitinib for the treatment of urothelial cancer, includingmetastatic urothelial cancer.

In one aspect of the present invention, a compound described herein canbe combined with at least one ERK inhibitor.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of SCH772984 forthe treatment of melanoma, including BRAF-mutant melanoma or NRAS-mutantmelanoma.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of ulixertinib forthe treatment of melanoma, including uveal melanoma.

In one embodiment, an effective amount of Compound 1 Pattern 1 isadministered in combination with an effective amount of ulixertinib forthe treatment of pancreatic cancer.

In one aspect of the present invention, a compound described herein canbe combined with at least one immunosuppressive agent. Theimmunosuppressive agent is preferably selected from the group consistingof a calcineurin inhibitor, e.g. a cyclosporin or an ascomycin, e.g.Cyclosporin A (NEORAL®), FK506 (tacrolimus), pimecrolimus, a mTORinhibitor, e.g. rapamycin or a derivative thereof, e.g. Sirolimus(RAPAMUNE®), Everolimus (Certican®), temsirolimus, zotarolimus,biolimus-7, biolimus-9, a rapalog, e.g., ridaforolimus, azathioprine,campath 1H, a SIP receptor modulator, e.g. fingolimod or an analoguethereof, an anti IL-8 antibody, mycophenolic acid or a salt thereof,e.g. sodium salt, or a prodrug thereof, e.g. Mycophenolate Mofetil(CELLCEPT®), OKT3 (ORTHOCLONE OKT3®), Prednisone, ATGAM®,THYMOGLOBULIN®, Brequinar Sodium, OKT4, T10B9.A-3A, 33B3.1,15-deoxyspergualin, tresperimus, Leflunomide ARAVA®, CTLAI-Ig,anti-CD25, anti-IL2R, Basiliximab (SIMULECT®), Daclizumab (ZENAPAX®),mizorbine, methotrexate, dexamethasone, ISAtx-247, SDZ ASM 981(pimecrolimus, Elidel®), CTLA41g (Abatacept), belatacept, LFA31g,etanercept (sold as Enbrel® by Immunex), adalimumab (Humira®),infliximab (Remicade®), an anti-LFA-1 antibody, natalizumab (Antegren®),Enlimomab, gavilimomab, antithymocyte immunoglobulin, siplizumab,Alefacept efalizumab, pentasa, mesalazine, asacol, codeine phosphate,benorylate, fenbufen, naprosyn, diclofenac, etodolac and indomethacin,aspirin and ibuprofen.

In certain embodiments, a compound described herein is administered tothe subject prior to treatment with another chemotherapeutic agent,during treatment with another chemotherapeutic agent, afteradministration of another chemotherapeutic agent, or a combinationthereof.

In some embodiments, an effective amount of the isolated Compound 1Pattern 1 can be administered to the subject such that the otherchemotherapeutic agent can be administered either at higher doses(increased chemotherapeutic dose intensity) or more frequently(increased chemotherapeutic dose density). Dose-dense chemotherapy is achemotherapy treatment plan in which drugs are given with less timebetween treatments than in a standard chemotherapy treatment plan.Chemotherapy dose intensity represents unit dose of chemotherapyadministered per unit time. Dose intensity can be increased or decreasedthrough altering dose administered, time interval of administration, orboth.

In one embodiment of the invention, the compounds described herein canbe administered in a concerted regimen with another agent such as anon-DNA-damaging, targeted anti-neoplastic agent or a hematopoieticgrowth factor agent. It has been recently been reported that theuntimely administration of hematopoietic growth factors can have seriousside effects. For example, the use of the EPO family of growth factorshas been associated with arterial hypertension, cerebral convulsions,hypertensive encephalopathy, thromboembolism, iron deficiency, influenzalike syndromes and venous thrombosis. The G-CSF family of growth factorshas been associated with spleen enlargement and rupture, respiratorydistress syndrome, allergic reactions and sickle cell complications. Assuch, in one embodiment, the use of the compounds or methods describedherein is combined with the use of hematopoietic growth factorsincluding, but not limited to, granulocyte colony stimulating factor(G-CSF, for example, sold as Neupogen (filgrastin), Neulasta(peg-filgrastin), or lenograstin), granulocyte-macrophage colonystimulating factor (GM-CSF, for example sold as molgramostim andsargramostim (Leukine)), M-CSF (macrophage colony stimulating factor),thrombopoietin (megakaryocyte growth development factor (MGDF), forexample sold as Romiplostim and Eltrombopag) interleukin (IL)-12,interleukin-3, interleukin-11 (adipogenesis inhibiting factor oroprelvekin), SCF (stem cell factor, steel factor, kit-ligand, or KL) anderythropoietin (EPO), and their derivatives (sold as for exampleepoetin-α as Darbopoetin, Epocept, Nanokine, Epofit, Epogin, Eprex andProcrit; epoetin-β sold as for example NeoRecormon, Recormon andMicera), epoetin-delta (sold as for example Dynepo), epoetin-omega (soldas for example Epomax), epoetin zeta (sold as for example Silapo andReacrit) as well as for example Epocept, EPOTrust, Erypro Safe,Repoeitin, Vintor, Epofit, Erykine, Wepox, Espogen, Relipoeitin,Shanpoietin, Zyrop and EPIAO). In one embodiment, an effective amount ofthe isolated Compound 1 Pattern 1 is administered prior toadministration of the hematopoietic growth factor. In one embodiment,the hematopoietic growth factor administration is timed so that thecompound's effect on HSPCs has dissipated. In one embodiment, the growthfactor is administered at least 20 hours after the administration of acompound described herein.

If desired, multiple doses of a compound described herein can beadministered to the subject. Alternatively, the subject can be given asingle dose of a compound described herein.

In one aspect of the invention, a compound disclosed herein can bebeneficially administered in combination with any therapeutic regimenentailing radiotherapy, chemotherapy, or other therapeutic agents. Inadditional embodiments the compounds disclosed herein can bebeneficially administered in combination with therapeutic agentstargeting auto-immune disorders.

EXAMPLES Example 1. X-Ray Powder Diffraction (XRPD)

XRPD analysis was carried out on a PANalytical X'pert pro, scanning thesamples between 3 and 35° 2θ. The material was gently grounded torelease any agglomerates and loaded onto a multi-well plate with Kaptonor Mylar polymer film to support the sample. The multi-well plate wasthen placed into the diffractometer and analyzed using Cu K radiation(α1 λ, =1.54060 Å; α2=1.54443 Å; β=1.39225 Å; α1: α2 ratio=0.5) runningin transmission mode (step size 0.0130° 2θ) using 40 kV/40 mA generatorsettings. The above technique was used to generate the images in FIG. 1,FIG. 7, FIG. 8 through FIG. 19, FIG. 21 through FIG. 32, FIG. 36, FIG.38, FIG. 42, FIG. 43, FIG. 49 through FIG. 59, FIG. 61, FIG. 67 throughFIG. 69, and FIG. 89.

Table 1 below provides the XRPD Peak List for Pattern 1 with >10%relative intensity peaks. The XRPD performed on Pattern 1 exhibitedsharp peaks, indicating the sample was composed of crystalline material.Significant peaks were observed in the XRPD on Pattern 1 at about9.6±0.2°, about 12.2±0.2°, about 15.3±0.2°, about 17.6±0.2°, about19.3±0.2°, about 19.8±0.2°, about 21.3±0.2°, about 22.7±0.2°, about24.0±0.2°, about 26.1±0.2°, and about 28.6±0.2°.

TABLE 1 XRPD Peak List for Pattern 1 Pos. [°2θ] d-spacing [Å] Rel. Int.[%] 9.57 9.24 100.0 10.12 8.74 16.80 12.20 7.26 57.10 13.82 6.41 14.3015.33 5.78 27.70 15.99 5.54 16.60 17.00 5.22 17.80 17.56 5.05 31.7018.71 4.74 11.50 19.28 4.60 35.70 19.84 4.47 65.40 20.30 4.37 23.3020.71 4.29 11.80 20.98 4.23 17.90 21.27 4.18 89.10 22.71 3.91 20.9023.19 3.84 24.70 23.97 3.71 53.30 26.06 3.42 37.00 26.76 3.33 16.4027.09 3.29 18.20 27.56 3.24 11.30 28.61 3.12 30.20 31.38 2.85 14.9032.30 2.77 13.60 32.87 2.72 13.00

In certain embodiments, the morphic form is Pattern 1 and ischaracterized by an XRPD pattern comprising at least 2 peaks selectedfrom about 9.6±0.2°, about 12.2±0.2°, about 15.3±0.2°, about 17.6±0.2°,about 19.3±0.2°, about 19.8±0.2°, about 21.2±0.2°, about 22.7±0.2°,about 23.9±0.2°, about 26.1±0.2°, and about 28.6±0.2°.

In certain embodiments, the morphic form is Pattern 1 and ischaracterized by an XRPD pattern comprising at least 3 peaks selectedfrom about 9.6±0.2°, about 12.2±0.2°, about 15.3±0.2°, about 17.6±0.2°,about 19.3±0.2°, about 19.8±0.2°, about 21.2±0.2°, about 22.7±0.2°,about 23.9±0.2°, about 26.1±0.2°, and about 28.6±0.2°.

In certain embodiments, the morphic form is Pattern 1 and ischaracterized by an XRPD pattern comprising at least 4 peaks selectedfrom about 9.6±0.2°, about 12.2±0.2°, about 15.3±0.2°, about 17.6±0.2°,about 19.3±0.2°, about 19.8±0.2°, about 21.2±0.2°, about 22.7±0.2°,about 23.9±0.2°, about 26.1±0.2°, and about 28.6±0.2°.

In certain embodiments, the morphic form is Pattern 1 and ischaracterized by an XRPD pattern comprising at least 5 peaks selectedfrom about 9.6±0.2°, about 12.2±0.2°, about 15.3±0.2°, about 17.6±0.2°,about 19.3±0.2°, about 19.8±0.2°, about 21.2±0.2°, about 22.7±0.2°,about 23.9±0.2°, about 26.1±0.2°, and about 28.6±0.2°.

In certain embodiments, the morphic form is Pattern 1 and ischaracterized by an XRPD pattern comprising at least 6 peaks selectedfrom about 9.6±0.2°, about 12.2±0.2°, about 15.3±0.2°, about 17.6±0.2°,about 19.3±0.2°, about 19.8±0.2°, about 21.2±0.2°, about 22.7±0.2°,about 23.9±0.2°, about 26.1±0.2°, and about 28.6±0.2°.

In certain embodiments, the morphic form is Pattern 1 and ischaracterized by an XRPD pattern comprising at least 7 peaks selectedfrom about 9.6±0.2°, about 12.2±0.2°, about 15.3±0.2°, about 17.6±0.2°,about 19.3±0.2°, about 19.8±0.2°, about 21.2±0.2°, about 22.7±0.2°,about 23.9±0.2°, about 26.1±0.2°, and about 28.6±0.2°.

In certain embodiments, the morphic form is Pattern 1 and ischaracterized by an XRPD pattern comprising at least 8 peaks selectedfrom about 9.6±0.2°, about 12.2±0.2°, about 15.3±0.2°, about 17.6±0.2°,about 19.3±0.2°, about 19.8±0.2°, about 21.2±0.2°, about 22.7±0.2°,about 23.9±0.2°, about 26.1±0.2°, and about 28.6±0.2°.

In certain embodiments, the morphic form is Pattern 1 and ischaracterized by an XRPD pattern comprising at least 9 peaks selectedfrom about 9.6±0.2°, about 12.2±0.2°, about 15.3±0.2°, about 17.6±0.2°,about 19.3±0.2°, about 19.8±0.2°, about 21.2±0.2°, about 22.7±0.2°,about 23.9±0.2°, about 26.1±0.2°, and about 28.6±0.2°.

In certain embodiments, the morphic form is Pattern 1 and ischaracterized by an XRPD pattern comprising at least 10 peaks selectedfrom about 9.6±0.2°, about 12.2±0.2°, about 15.3±0.2°, about 17.6±0.2°,about 19.3±0.2°, about 19.8±0.2°, about 21.2±0.2°, about 22.7±0.2°,about 23.9±0.2°, about 26.1±0.2°, and about 28.6±0.2°.

In certain embodiments, the morphic form is Pattern 1 and ischaracterized by an XRPD pattern comprising the 2theta values selectedfrom about 9.6±0.2°, about 12.2±0.2°, about 15.3±0.2°, about 17.6±0.2°,about 19.3±0.2°, about 19.8±0.2°, about 21.2±0.2°, about 22.7±0.2°,about 23.9±0.2°, about 26.1±0.2°, and about 28.6±0.2°.

Table 2 below provides the results of the XRPD performed on Pattern 2.The XRPD exhibited sharp peaks, indicating the sample was composed ofcrystalline material. Significant peaks were observed in the XRPD onPattern 2 at 6.7±0.2°, 11.1±0.2°, 16.3±0.2°, 17.2±0.2°, 18.2±0.2°,19.8±0.2°, 20.4±0.2°, 20.6±0.2°, 26.4±0.2°, and 27.3±0.2°.

TABLE 2 XRPD peaks of Pattern 2. Pos. [°2θ] d-spacing [Å] Rel. Int. [%]6.77 13.050 100 11.15 7.933 58.67 14.06 6.300 14.44 14.77 5.996 24.2115.08 5.875 19.26 16.29 5.442 63.87 16.96 5.227 26.73 17.24 5.144 36.6218.19 4.878 39.51 19.81 4.482 36.17 20.42 4.350 33.11 20.64 4.303 31.3621.55 4.125 23.41 22.72 3.914 22.54 23.11 3.849 28.74 24.90 3.576 16.5525.55 3.486 14.30 26.41 3.375 99.54 27.28 3.269 49.45 28.46 3.137 27.8530.35 2.945 10.69

In certain embodiments, the morphic form is Pattern 2 and ischaracterized by an XRPD pattern comprising at least 2 peaks selectedfrom 6.7±0.2°, 11.1±0.2°, 16.3±0.2°, 17.2±0.2°, 18.2±0.2°, 19.8±0.2°,20.4±0.2°, 20.6±0.2°, 26.4±0.2°, and 27.3±0.2°.

In certain embodiments, the morphic form is Pattern 2 and ischaracterized by an XRPD pattern comprising at least 3 peaks selectedfrom 6.7±0.2°, 11.1±0.2°, 16.3±0.2°, 17.2±0.2°, 18.2±0.2°, 19.8±0.2°,20.4±0.2°, 20.6±0.2°, 26.4±0.2°, and 27.3±0.2°.

In certain embodiments, the morphic form is Pattern 2 and ischaracterized by an XRPD pattern comprising at least 4 peaks selectedfrom 6.7±0.2°, 11.1±0.2°, 16.3±0.2°, 17.2±0.2°, 18.2±0.2°, 19.8±0.2°,20.4±0.2°, 20.6±0.2°, 26.4±0.2°, and 27.3±0.2°.

In certain embodiments, the morphic form is Pattern 2 and ischaracterized by an XRPD pattern comprising at least 5 peaks selectedfrom 6.7±0.2°, 11.1±0.2°, 16.3±0.2°, 17.2±0.2°, 18.2±0.2°, 19.8±0.2°,20.4±0.2°, 20.6±0.2°, 26.4±0.2°, and 27.3±0.2°.

In certain embodiments, the morphic form is Pattern 2 and ischaracterized by an XRPD pattern comprising at least 6 peaks selectedfrom 6.7±0.2°, 11.1±0.2°, 16.3±0.2°, 17.2±0.2°, 18.2±0.2°, 19.8±0.2°,20.4±0.2°, 20.6±0.2°, 26.4±0.2°, and 27.3±0.2°.

In certain embodiments, the morphic form is Pattern 2 and ischaracterized by an XRPD pattern comprising at least 7 peaks selectedfrom 6.7±0.2°, 11.1±0.2°, 16.3±0.2°, 17.2±0.2°, 18.2±0.2°, 19.8±0.2°,20.4±0.2°, 20.6±0.2°, 26.4±0.2°, and 27.3±0.2°.

In certain embodiments, the morphic form is Pattern 2 and ischaracterized by an XRPD pattern comprising at least 8 peaks selectedfrom 6.7±0.2°, 11.1±0.2°, 16.3±0.2°, 17.2±0.2°, 18.2±0.2°, 19.8±0.2°,20.4±0.2°, 20.6±0.2°, 26.4±0.2°, and 27.3±0.2°.

In certain embodiments, the morphic form is Pattern 2 and ischaracterized by an XRPD pattern comprising at least 9 peaks selectedfrom 6.7±0.2°, 11.1±0.2°, 16.3±0.2°, 17.2±0.2°, 18.2±0.2°, 19.8±0.2°,20.4±0.2°, 20.6±0.2°, 26.4±0.2°, and 27.3±0.2°.

In certain embodiments, the morphic form is Pattern 2 and ischaracterized by an XRPD pattern comprising the 2theta values selectedfrom 6.7±0.2°, 11.1±0.2°, 16.3±0.2°, 17.2±0.2°, 18.2±0.2°, 19.8±0.2°,20.4±0.2°, 20.6±0.2°, 26.4±0.2°, and 27.3±0.2°.

Table 4 below provides the results of the XRPD performed on Pattern 5.Significant peaks were observed in the XRPD on Pattern 5 at 6.3±0.2°,6.8±0.2°, 8.1±0.2°, 11.2±0.2°, 16.2±0.2°, 26.5±0.2°, 27.4±0.2°,27.8±0.2°, and 28.6±0.2°.

Table 2 below provides the results of the XRPD performed on Pattern 3scale-up. The XRPD exhibited sharp peaks, indicating the sample wascomposed of crystalline material. Significant peaks were observed in theXRPD on Pattern 3 scale-up at about 8.1±0.2°, about 14.8±0.2°, about16.5±0.2°, about 18.2±0.2°, about 19.7±0.2°, about 24.8±0.2°, about25.6±0.2° and about 27.7±0.2°.

TABLE 3 XRPD peaks for Pattern 3. Pos. [°2θ] d-spacing [Å] Rel. Int. [%]8.12 10.885 100 16.50 5.371 59.88 24.84 3.584 40.08 14.80 5.986 37.3618.16 4.884 33.40 27.72 3.218 31.65 25.56 3.485 28.53 19.74 4.497 27.6117.13 5.177 27.38 12.99 6.817 26.27 15.98 5.548 25.23 22.20 4.005 21.0426.76 3.331 19.56 24.30 3.662 19.00 31.30 2.858 18.31 5.13 17.212 17.6429.87 2.992 13.67 33.15 2.703 11.17 29.06 3.073 10.28

In certain embodiments, the morphic form is Pattern 3 is characterizedby an XRPD pattern comprising at least 2 peaks selected from 8.1±0.2°,14.8±0.2°, 16.5±0.2°, 18.1±0.2°, 19.7±0.2°, 24.84±0.2°, and 27.7±0.2°.

In certain embodiments, the morphic form is Pattern 3 is characterizedby an XRPD pattern comprising at least 3 peaks selected from 8.1±0.2°,14.8±0.2°, 16.5±0.2°, 18.1±0.2°, 19.7±0.2°, 24.84±0.2°, and 27.7±0.2°.

In certain embodiments, the morphic form is Pattern 3 is characterizedby an XRPD pattern comprising at least 4 peaks selected from 8.1±0.2°,14.8±0.2°, 16.5±0.2°, 18.1±0.2°, 19.7±0.2°, 24.84±0.2°, and 27.7±0.2°.

In certain embodiments, the morphic form is Pattern 3 is characterizedby an XRPD pattern comprising at least 5 peaks selected from 8.1±0.2°,14.8±0.2°, 16.5±0.2°, 18.1±0.2°, 19.7±0.2°, 24.84±0.2°, and 27.7±0.2°.

In certain embodiments, the morphic form is Pattern 3 is characterizedby an XRPD pattern comprising at least 6 peaks selected from 8.1±0.2°,14.8±0.2°, 16.5±0.2°, 18.1±0.2°, 19.7±0.2°, 24.84±0.2°, and 27.7±0.2°.

In certain embodiments, the morphic form is Pattern 3 is characterizedby an XRPD pattern comprising the 2theta values selected from 8.1±0.2°,14.8±0.2°, 16.5±0.2°, 18.1±0.2°, 19.7±0.2°, 24.84±0.2°, and 27.7±0.2°.

TABLE 4 XRPD peaks of Pattern 4. Pos. [°2θ] d-spacing [Å] Rel. Int. [%]6.36 13.896 35.99 6.79 13.009 95.80 8.11 10.903 41.76 11.21 7.892 49.5113.59 6.518 13.93 14.12 6.274 16.30 14.87 5.960 23.31 15.16 5.843 23.6116.24 5.457 70.61 17.30 5.125 31.55 18.27 4.855 36.30 18.98 4.677 17.4719.89 4.464 28.00 20.49 4.334 26.21 20.72 4.288 27.97 21.65 4.105 17.9422.82 3.897 17.26 23.21 3.833 24.78 24.76 3.595 19.75 25.63 3.476 17.1226.53 3.360 100 27.41 3.254 56.04 27.85 3.203 37.88 28.61 3.120 41.9429.39 3.039 12.96 30.02 2.977 13.48 30.42 2.939 16.39 31.42 2.847 10.7532.50 2.755 13.49 33.07 2.709 12.96

In certain embodiments, the morphic form is Pattern 5 and ischaracterized by an XRPD pattern comprising at least 2 peaks selectedfrom 6.3±0.2°, 6.8±0.2°, 8.1±0.2°, 11.2±0.2°, 16.2±0.2°, 26.5±0.2°,27.4±0.2°, 27.8±0.2°, and 28.6±0.2°.

In certain embodiments, the morphic form is Pattern 5 and ischaracterized by an XRPD pattern comprising at least 3 peaks selectedfrom 6.3±0.2°, 6.8±0.2°, 8.1±0.2°, 11.2±0.2°, 16.2±0.2°, 26.5±0.2°,27.4±0.2°, 27.8±0.2°, and 28.6±0.2°.

In certain embodiments, the morphic form is Pattern 5 and ischaracterized by an XRPD pattern comprising at least 4 peaks selectedfrom 6.3±0.2°, 6.8±0.2°, 8.1±0.2°, 11.2±0.2°, 16.2±0.2°, 26.5±0.2°,27.4±0.2°, 27.8±0.2°, and 28.6±0.2°.

In certain embodiments, the morphic form is Pattern 5 and ischaracterized by an XRPD pattern comprising at least 5 peaks selectedfrom 6.3±0.2°, 6.8±0.2°, 8.1±0.2°, 11.2±0.2°, 16.2±0.2°, 26.5±0.2°,27.4±0.2°, 27.8±0.2°, and 28.6±0.2°.

In certain embodiments, the morphic form is Pattern 5 and ischaracterized by an XRPD pattern comprising at least 6 peaks selectedfrom 6.3±0.2°, 6.8±0.2°, 8.1±0.2°, 11.2±0.2°, 16.2±0.2°, 26.5±0.2°,27.4±0.2°, 27.8±0.2°, and 28.6±0.2°.

In certain embodiments, the morphic form is Pattern 5 and ischaracterized by an XRPD pattern comprising at least 7 peaks selectedfrom 6.3±0.2°, 6.8±0.2°, 8.1±0.2°, 11.2±0.2°, 16.2±0.2°, 26.5±0.2°,27.4±0.2°, 27.8±0.2°, and 28.6±0.2°.

In certain embodiments, the morphic form is Pattern 5 and ischaracterized by an XRPD pattern comprising at least 8 peaks selectedfrom 6.3±0.2°, 6.8±0.2°, 8.1±0.2°, 11.2±0.2°, 16.2±0.2°, 26.5±0.2°,27.4±0.2°, 27.8±0.2°, and 28.6±0.2°.

In certain embodiments, the morphic form is Pattern 5 and ischaracterized by an XRPD pattern comprising the 2theta values selectedfrom 6.3±0.2°, 6.8±0.2°, 8.1±0.2°, 11.2±0.2°, 16.2±0.2°, 26.5±0.2°,27.4±0.2°, 27.8±0.2°, and 28.6±0.2°.

\ Table 5 below provides the results of the XRPD performed on Pattern 6exhibited sharp peaks, indicating the sample was composed of crystallinematerial. Tabulated peak picks for Pattern 6>10% relative intensity.Significant peaks were observed in the XRPD on Pattern 6 at 6.7±0.2°,11.1±0.2°, 16.26±0.2°, 26.3±0.2°, and 27.2±0.2°.

TABLE 5 XRPD peaks for Pattern 6 scale-up. Pos. [°2θ] d-spacing [Å] Rel.Int. [%] 6.77 13.051 100 9.54 9.273 15.73 11.15 7.938 55.43 14.74 6.01212.56 16.26 5.452 55.44 16.94 5.234 21.60 17.23 5.146 32.11 18.16 4.88533.45 19.78 4.490 33.56 20.39 4.355 24.11 20.62 4.308 23.52 21.20 4.19218.54 21.51 4.131 14.66 22.68 3.920 14.28 23.08 3.853 27.52 23.91 3.72112.10 24.85 3.582 11.69 26.02 3.424 20.04 26.36 3.381 83.70 27.26 3.27142.92 28.43 3.140 27.55

In certain embodiments, the morphic form is Pattern 6 and ischaracterized by an XRPD pattern comprising at least 2 peaks selectedfrom 6.7±0.2°, 11.1±0.2°, 16.26±0.2°, 26.3±0.2°, and 27.2±0.2°.

In certain embodiments, the morphic form is Pattern 6 and ischaracterized by an XRPD pattern comprising at least 3 peaks selectedfrom 6.7±0.2°, 11.1±0.2°, 16.26±0.2°, 26.3±0.2°, and 27.2±0.2°.

In certain embodiments, the morphic form is Pattern 6 and ischaracterized by an XRPD pattern comprising at least 4 peaks selectedfrom 6.7±0.2°, 11.1±0.2°, 16.26±0.2°, 26.3±0.2°, and 27.2±0.2°.

In certain embodiments, the morphic form is Pattern 6 and ischaracterized by an XRPD pattern comprising the 2theta values selectedfrom 6.7±0.2°, 11.1±0.2°, 16.26±0.2°, 26.3±0.2°, and 27.2±0.2°.

Example 2. General Technique for using Gravimetric Vapor Sorption (GVS)

Approximately 10-20 mg of sample was placed into a mesh vapor sorptionbalance pan and loaded into an IGASorp Moisture Sorption Analyzerbalance by Hiden Analytical. The sample was subjected to a rampingprofile from 40-90% relative humidity (RH) at 10% increments,maintaining the sample at each step until a stable weight had beenachieved (98% step completion, minimum step length 30 minutes, maximumstep length 60 minutes) at 25° C. After completion of the sorptioncycle, the sample was dried using the same procedure to 0% RH, andfinally taken back to the starting point of 40% RH. Three cycles wereperformed. The weight change during the sorption/desorption cycles wereplotted, allowing for the hygroscopic nature of the sample to bedetermined. The above technique was used to generate the images in FIG.40 and FIG. 41.

Table 6 below provides tabulated GVS data for Pattern 1 showing thedifference in water vapor uptake between the sorption and desorptionisotherms. The weight change during the sorption/desorption cycles wereplotted, allowing for the hygroscopic nature of the sample to bedetermined.

TABLE 6 Tabulated GVS data for Pattern 1 Target Change in Mass (%) - ref(RH (%) Sorption Desorption Cycle 1 0 0.180 10 2.320 20 2.820 30 3.11040 3.420 3.340 50 3.560 3.580 60 3.720 3.730 70 3.800 3.820 80 3.9203.970 90 4.160 4.160 Cycle 2 0 0.180 0.000 10 2.300 2.240 20 2.770 2.68030 3.030 3.020 40 3.290 3.260 50 3.470 3.500 60 3.660 3.670 70 3.7803.780 80 3.900 3.890 90 4.080 4.080 Cycle 3 0.000 0.000 10.000 2.25020.000 2.740 30.000 3.020 40.000 3.220

In certain embodiments, the sample was subjected to a ramping profilefrom 40-90% relative humidity (RH) at 10% increments.

In certain embodiments, at least two sorption cycles are performed.

In certain embodiments, three sorption cycles are performed.

Table 7 below provides the results of the GVS data for the mixture ofPattern 2 and pattern 6, showing the difference in water vapor uptakebetween the sorption and desorption isotherms. The weight change duringthe sorption/desorption cycles were plotted, allowing for thehygroscopic nature of the sample to be determined.

TABLE 7 GVS data of Pattern 2 and Pattern 6 mixture Target Change inMass (%) - ref (RH (%) Sorption Desorption Cycle 1 0 0.000 10 3.118 203.734 30 4.148 40 4.737 4.497 50 5.420 4.806 60 7.044 5.181 70 6.3465.653 80 6.770 6.343 90 7.898 7.898 Cycle 2 0 0.000 0.059 10 2.731 3.20020 3.470 3.820 30 3.969 4.205 40 4.353 4.512 50 4.661 4.848 60 4.9995.157 70 5.436 5.572 80 5.976 6.181 90 7.039 7.039 Cycle 3 0 0.059 102.708 20 3.502 30 4.027 40 4.335

In certain embodiments, the sample was subjected to a ramping profilefrom 40-90% relative humidity (RH) at 10% increments.

In certain embodiments, at least two sorption cycles are performed.

In certain embodiments, three sorption cycles are performed.

Example 3. Dynamic Vapor Sorption (DVS)

Approximately, 10-20 mg of sample was placed into a mesh vapor sorptionbalance pan and loaded into a DVS Intrinsic dynamic vapor sorptionbalance by Surface Measurement Systems. The sample was subjected to aramping profile from 40-90% relative humidity (RH) at 10% increments,maintaining the sample at each step until a stable weight had beenachieved (dm/dt 0.004%, minimum step length 30 minutes, maximum steplength 500 minutes) at 25° C. After completion of the sorption cycle,the sample was dried using the same procedure to 0% RH and then a secondsorption cycle back to 40% RH. Two cycles were performed. The weightchange during the sorption/desorption cycles were plotted, allowing forthe hygroscopic nature of the sample to be determined. XRPD analysis wasthen carried out on any solid retained. The above technique was used togenerate the images in FIG. 5, FIG. 6, FIG. 47 and FIG. 48.

TABLE 8 provides results for the tabulated DVS data of Pattern 3scale-up. Target Change in Mass (%) - ref (RH (%) Sorption DesorptionHysteresis Cycle 1 0 0.000 10 0.863 20 1.274 30 1.818 40 2.516 2.223−0.292 50 2.780 2.491 −0.289 60 2.970 2.727 −0.242 70 3.115 2.940 −0.17580 3.288 3.171 −0.117 90 3.555 3.555 Cycle 2 0 0.000 0.075 10 0.8310.885 0.054 20 1.198 1.290 0.091 30 1.468 1.711 0.243 40 1.853 1.9770.124 50 2.092 2.171 0.080 60 2.271 2.337 0.066 70 2.461 2.510 0.050 802.700 2.717 0.018 90 3.093 3.093 Cycle 3 0 0.075 10 0.861 20 1.198 301.443 40 1.757

The material appeared hygroscopic by DVS with a mass increase of ca. 4%at 90% RH. During the desorption cycles the material loses mass, likelydue to the loss of surface water present in the sample and thecrystallization of amorphous content present in the sample. The materialadsorbed 2.5 wt % moisture at 40% RH, 3.1 wt % moisture at 60% and 3.5wt % at 90% RH.

In certain embodiments, at least two sorption cycles are performed. Incertain embodiments, three sorption cycles are performed.

Example 4. Tabulated Peak Picks of Pattern 4>5% Relative Intensity

XRPD analysis was carried out on a PANalytical X'pert pro, scanning thesamples between 3 and 35° 2θ. The material was gently ground to releaseany agglomerates and loaded onto a multi-well plate with Kapton or Mylarpolymer film to support the sample. The multi-well plate was then placedinto the diffractometer and analyzed using Cu K radiation (α1 λ,=1.54060 Å; α2=1.54443 Å; β=1.39225 Å; α1:α2 ratio=0.5) running intransmission mode (step size 0.0130° 2θ) using 40 kV/40 mA generatorsettings. The above technique was used to generate the images in FIG.13, FIG. 20 and FIG. 27.

Table 9 below provides the results of the XRPD performed on Pattern 4exhibited sharp peaks, indicating the sample was composed of crystallinematerial. Significant peaks were observed in the XRPD on Pattern 4 at6.1±0.2°, 15.0±0.2°, 27.5±0.2°, and 28.1±0.2°.

TABLE 9 XRPD peaks for Pattern 4 Pos. [°2θ] d-spacing [Å] Rel. Int. [%]6.13 14.416 100 9.55 9.265 20.37 14.64 6.049 14.50 15.03 5.896 21.2917.84 4.971 12.89 18.44 4.811 12.04 20.24 4.387 6.94 21.25 4.181 10.2221.97 4.046 5.55 25.73 3.462 6.79 26.42 3.374 6.40 27.55 3.238 29.6028.15 3.170 29.11 29.61 3.017 6.00

In certain embodiments, the morphic form is Pattern 4 and ischaracterized by an XRPD pattern comprising at least 2 peaks selectedfrom 6.1±0.2°, 15.0±0.2°, 27.5±0.2°, and 28.1±0.2 °.

In certain embodiments, the morphic form is Pattern 4 and ischaracterized by an XRPD pattern comprising at least 3 peaks selectedfrom 6.1±0.2°, 15.0±0.2°, 27.5±0.2°, and 28.1±0.2°.

In certain embodiments, the morphic form is Pattern 4 and ischaracterized by an XRPD pattern comprising the 2theta values selectedfrom 6.1±0.2°, 15.0±0.2°, 27.5±0.2°, and 28.1±0.2°.

Example 5. PLM Imaging

PLM imaging was carried out in the presence of crystallinity(birefringence). PLM imaging was determined using an Olympus BX50polarizing microscope, equipped with a Motic camera and image capturesoftware (Motic Images Plus 2.0). All images were recorded using the 20×objective, unless otherwise stated. The above technique was used togenerate the images in FIG. 2, FIG. 37, FIG. 44, FIG. 62, and FIG. 74through FIG. 86.

Example 6. Thermogravimetric Analysis (TGA)

Approximately 5 mg of material was weighed into an open aluminum pan andloaded into a simultaneous thermogravimetric/differential thermalanalyzer (TG/DTA) and held at room temperature. The sample was thenheated at a rate of 10° C./min from 20° C. to 300° C. during which timethe change in sample weight was recorded along with any differentialthermal events (DTA). Nitrogen was used as the purge gas, at a flow rateof 300 cm³/min. The above technique was used to generate the images inFIG. 3, FIG. 33, FIG. 34, FIG. 35, FIG. 38, FIG. 45, FIG. 63, FIG. 87and FIG. 88.

Example 7. Differential Scanning Calorimetry (DSC)

Approximately, 5 mg of material was weighed into an aluminum DSC pan andsealed non-hermetically with a pierced aluminum lid. The sample pan wasthen loaded into a Seiko DSC6200 (equipped with a cooler) cooled andheld at 20° C. Once a stable heat-flow response was obtained, the sampleand reference were heated to 360° C. at a scan rate of 10° C./min andthe resulting heat flow response monitored. Nitrogen was used as thepurge gas, at a flow rate of 50 cm³/min. The above technique was used togenerate the images in FIG. 4, FIG. 39, FIG. 46, FIG. 64, FIG. 65 andFIG. 66.

Example 8. Crystallization and Competitive Slurry Experiments

Competitive slurry experiments were carried out using ca. 10 mg ofPattern 1, Pattern 2/6 mixture and Pattern 3 were weighed out and addedto a 1.5 mL glass vial. Samples were prepared using 100 μL aliquots ofsolvent until a mobile slurry was formed. Samples were stirred atambient temperature for 48 hours. A second set of samples were stirredat 60° C. for 48 hours. All samples were collected, and the solids wereisolated and analyzed by XRPD. The above technique was used to generatethe images in FIG. 50, FIG. 51, and FIG. 55 through FIG. 59.

TABLE 10 Results and observations from competitive slurry experimentsVolume Input of Solvent Sample Material Solvent (μL) TemperatureObservations XRPD Form 1 1 + 2 + 3 EtOH 1000 Ambient Yellow 1 slurry 21 + 2 + 3 MEK 1000 Ambient Yellow 1 with traces of 2 slurry 3 1 + 2 + 32N HCl 1000 Ambient Yellow 1 slurry 4 1 + 2 + 3 IPA 1000 Ambient Yellow1 with traces of 2 slurry 5 1 + 2 + 3 MeCN/ 1000 Ambient Yellow 1 waterslurry (97:3% v/v) 6 1 + 2 + 3 Acetone/ 1000 Ambient Yellow 1 waterslurry (85:15% v/v) 7 1 + 2 + 3 EtOH 1000 60° C. Yellow 1 slurry 8 1 +2 + 3 MEK 1000 60° C. Yellow Mix of 1, 2 and 3 slurry 9 1 + 2 + 3 2N HCl1000 60° C. Yellow 1 with traces of 2 slurry 10 1 + 2 + 3 IPA 1000 60°C. Yellow 1 with traces of 2 slurry 11 1 + 2 + 3 MeCN/ 1000 60° C.Yellow 1 water slurry (97:3% v/v) 12 1 + 2 + 3 Acetone/ 1000 60° C.Yellow 1 water slurry (85:15% v/v)

Example 9. XRPD Analysis after DVS Experiment

XRPD analysis was carried out on a PANalytical X'pert pro, scanning thesamples between 3 and 35° 2θ. The material was gently grounded torelease any agglomerates and loaded onto a multi-well plate with Kaptonor Mylar polymer film to support the sample. The multi-well plate wasthen placed into the diffractometer and analyzed using Cu K radiation(α1 λ=1.54060 Å; α2 =1.54443 Å; β=1.39225 Å; α1:α2 ratio=0.5) running intransmission mode (step size 0.0130° 2θ) using 40 kV/40 mA generatorsettings.

The post-DVS samples were analyzed by XRPD. After DVS was performed,samples were analyzed by XRPD to confirm any changes in the material.The above technique was used to generate the images in FIG. 7, FIG. 42and FIG. 49.

Example 10. Stability Study after 1 Week

Stability study was carried out using ca. 10 mg of material. Sampleswere prepared in duplicate for XRPD and HPLC analysis. One set ofsamples were capped and stored under ambient light, temperature andhumidity. One set of samples were capped and stored in an oven at 80° C.One set of samples (75%) were uncapped and stored in a desiccatorcontaining sodium chloride solution at 40° C. After 7 days the sampleswere collected and analyzed by XRPD and HPLC. The above technique wasused to generate the images in FIG. 52, FIG. 53 and FIG. 54.

Example 11. Various Solvent Solubility Systems

The solvent solubility experiment was carried out using various solventsto determine solid formation. A known volume aliquot (typically 5volumes) of solvent was added to approximately 10 mg of amorphousCompound 1 as a dihydrochloride. Between each addition, the mixture waschecked for dissolution and where no dissolution was apparent, themixture was heated to ca. 40° C. and checked again. This procedure wascontinued until dissolution was observed or until 100 volumes of solventhad been added. All observed solids were isolated and analyzed by XRPD.

Examples of solvents include, but are not limited to, 1,4 dioxane,methanol, ethanol, propanol, butanol, butanone, ethoxyethanol,trifluoroethanol, methyl-THF, acetone, acetonitrile, anisole,chlorobenzene, DMSO, DMF, cumene, dichloromethane, ethyl acetate,ethylene glycol, heptane, isopropyl acetate, nitromethane,n-methylpyrrolidone, MIBK, t-BME, tetrahydrofuran, toluene, water andmixtures thereof. The above technique was used to generate the images inFIG. 8, FIG. 9, FIG. 10, FIG. 11, FIG. 12, FIG. 14.

TABLE 11 Approximate solubility results from solvent screen Approx.Solubility Sample Solvent (mg/mL) 1 1,4-Dioxane <5 2 1-Butanol <5 32,2,2-Trifluoroethanol >200 4 2-Butanone <5 5 2-Ethoxyethanol <5 62-Methyl THF <5 7 2-Propanol ca 5 8 Acetone <5 9 Acetonitrile <5 10Acetonitrile/water (90:10% v/v) <5 11 Anisole <5 12 Chlorobenzene <5 13DMSO ca 5 14 DMF <5 15 Cumene ca 5 16 Dichloromethane <5 17 Ethanol ca 518 Ethanol/water (85:15% v/v) <5 19 Ethyl Acetate <5 20 Ethylene glycol50 21 Heptane <5 22 Isopropyl acetate <5 23 Methanol ca 5 24Methanol/water (95:5% v/v) 20 25 Nitromethane 33 26 N-Methylpyrrolidone<5 27 Methyl isobutyl ketone <5 28 t-Butylmethyl ether <5 29Tetrahydrofuran <5 30 Tetrahydrofuran/water (99:1% v/v) <5 31 Toluene <532 Deionized Water >200

TABLE 12 XRPD results from solvent solubility screen Sample Solvent XRPD1 1,4-Dioxane Pattern 1 2 1-Butanol Pattern 2 3 2,2,2-TrifluoroethanolPattern 3 4 2-Butanone Pattern 2 5 2-Ethoxyethanol Pattern 2 6 2-MethylTHF Pattern 2 7 2-Propanol Pattern 2 8 Acetone Pattern 5 9 AcetonitrilePattern 4 10 Acetonitrile/water (90:10% v/v) Pattern 1 11 AnisolePattern 6 12 Chlorobenzene Amorphous 13 DMSO Amorphous 14 DMF Amorphous15 Cumene Pattern 2 16 Dichloromethane Pattern 1 17 Ethanol Pattern 1 18Ethanol/water (85:15% v/v) Pattern 1 19 Ethyl Acetate Pattern 2 20Ethylene glycol No solid 21 Heptane Pattern 3 22 Isopropyl acetatePattern 2 23 Methanol Pattern 1 24 Methanol/water (95:5% v/v) Pattern 125 Nitromethane Pattern 1 26 N-Methylpyrrolidone Pattern 2 27 Methylisobutyl ketone Pattern 2 28 t-Butylmethyl ether Pattern 5 29Tetrahydrofuran Pattern 1 30 Tetrahydrofuran/water (99:1% v/v) Pattern 131 Toluene Pattern 2 32 Water No solid

After removing the initial solid, samples in which dissolution wasobserved were uncapped and allowed to evaporate. This resulted in thediffractograms in FIG. 30 and FIG. 31.

TABLE 13 Observations from the evaporation experiments ObservationsPost- Sample Solvent System Evaporation 1 1,4-Dioxane No solid 2Acetone/water (20:80% v/v) Yellow solid 3 2,2,2-Trifluoroethanol Yellowsolid 4 2-Butanone No solid 5 2-Propanol No solid 6 Acetone No solid 7Acetonitrile No solid 8 Acetonitrile/water (90:10% v/v) No solid 9Dichloromethane No solid 10 Ethanol Small amount of yellow solid 11Ethanol/water (85:15% v/v) Small amount of yellow solid 12 Ethyl AcetateNo solid 13 Heptane No solid 14 Isopropyl acetate No solid 15 MethanolNo solid 16 Methanol/water (95:5% v/v) Small amount of yellow solid 172N HCl No solid 18 Methyl isobutyl ketone No solid 19 t-Butylmethylether No solid 20 Tetrahydrofuran No solid 21 Toluene No solid

Example 12. Anti-Solvent Addition Experiments

The antisolvent addition experiments were carried out using selectedanti-solvent (for example t-BME) layered over the material solution ineach sample until precipitation was observed or 1 mL of anti-solvent wasadded. The samples were stored in a fridge at ca. 5° C. to promoteprecipitation and any observed solids were collected and analyzed byXRFD. Material solution were prepared using solvent systems listed inExample 13. The above technique was used to generate the images in FIG.32.

TABLE 14 Observations from the anti-solvent addition experiments Volumeof Anti- Sam- Anti- Solvent Observations Post AS ple Solvent SystemSolvent (μL) Addition 1 1,4-Dioxane t-BME 1000 Clear colorless solution2 Acetone/water t-BME 1000 Clear colorless solution (20:80% v/v) 32,2,2- t-BME 500 Yellow solid Trifluoroethanol 4 2-Butanone t-BME 1000Clear colorless solution 5 2-Ethoxyethanol t-BME 1000 Clear colorlesssolution 6 2-Propanol t-BME 1000 Clear colorless solution 7 Acetonet-BME 1000 Clear colorless solution 8 Acetonitrile t-BME 1000 Clearcolorless solution 9 Acetonitrile/water t-BME 1000 Clear colorlesssolution (90:10% v/v) 10 Anisole t-BME 1000 Clear colorless solution 11Cumene t-BME 1000 Clear colorless solution 12 Dichloromethane t-BME 1000Clear colorless solution 13 Ethanol t-BME 1000 Clear colorless solution14 Ethanol/water t-BME 1000 Clear colorless solution (85:15% v/v) 15Ethyl Acetate t-BME 1000 Clear colorless solution 16 Heptane t-BME 1000Clear colorless solution 17 Isopropyl acetate t-BME 1000 Clear colorlesssolution 18 Methanol t-BME 1000 Clear colorless solution 19Methanol/water t-BME 1000 Clear colorless solution (95:5% v/v) 20 2N HClt-BME 1000 Clear colorless solution 21 Methyl isobutyl t-BME 1000 Clearcolorless solution ketone 22 t-Butylmethyl Heptane 1000 Clear colorlesssolution ether 23 Tetrahydrofuran t-BME 1000 Clear colorless solution 24Toluene t-BME 1000 Clear colorless solution

TABLE 15 Observed solids from the anti-solvent addition experimentsAnti- Sample Solvent System Solvent XRPD 1 1,4-Dioxane t-BME No solid 2Acetone/water (20:80% v/v) t-BME No solid 3 2,2,2-Trifluoroethanol t-BMEPattern 3 4 2-Butanone t-BME No solid 5 2-Ethoxyethanol t-BME No solid 62-Propanol t-BME No solid 7 Acetone t-BME No solid 8 Acetonitrile t-BMENo solid 9 Acetonitrile/water (90:10% t-BME No solid v/v) 10 Anisolet-BME No solid 11 Cumene t-BME No solid 12 Dichloromethane t-BME Nosolid 13 Ethanol t-BME No solid 14 Ethanol/water (85:15% v/v) t-BME Nosolid 15 Ethyl Acetate t-BME No solid 16 Heptane t-BME No solid 17Isopropyl acetate t-BME No solid 18 Methanol t-BME No solid 19Methanol/water (95:5% v/v) t-BME No solid 20 2N HCl t-BME No solid 21Methyl isobutyl ketone t-BME No solid 22 t-Butylmethyl ether Heptane Nosolid 23 Tetrahydrofuran t-BME No solid 24 Toluene t-BME No solid

Example 13. Scale Up of Pattern 2, Pattern 3, and Pattern 5

Pattern 2 was scaled up via the addition of 100 uL aliquots of MEK toapproximately 500 mg of amorphous Compound 1 as a di HCl salt until amobile slurry was formed. The slurry was then temperature cycled betweenambient and 40° C. for approximately 72 hours. The sample was collected,and an aliquot of the slurry was analyzed by XRPD. The solid was thenisolated and dried under vacuum at 40° C. for approximately 4 hours. Theresulting material was a mixture of Pattern 2 and Pattern 6.

Pattern 3 was scaled up via the addition of 100 uL aliquots of ethanolto approximately 500 mg of amorphous Compound 1 as a di HCl salt until amobile slurry was formed. The slurry was then temperature cycled betweenambient and 40° C. for approximately 72 hours. The sample was collected,and an aliquot of the slurry was analyzed by XRPD. The solid was thenisolated and dried under vacuum at 40° C. for approximately 4 hours.

Pattern 5 was scaled up via the addition of 100 μL aliquots of ethanolto approximately 500 mg of amorphous Compound 1 as a di HCl salt until amobile slurry was formed. The slurry was then temperature cycled betweenambient and 40° C. for approximately 72 hours. The sample was collected,and an aliquot of the slurry was analyzed by XRPD. The solid was thenisolated and dried under vacuum at 40° C. for approximately 4 hours.This resulting material was then slurried in 10 mL of methyl isobutylketone for another twenty-four hours. An aliquot of the slurry wasanalyzed by XRPD. The solid was then isolated and dried under vacuum at40° C. for approximately four hours. The dried material was analyzed byXRPD.

Example 14. The pH Solubility and pH Solvent Systems

Experiments of pH solubility were carried out on Pattern 7 solids at pH2 to pH 13. A total of 12 vials contained Pattern 7 (100 mg) and 500 μLof 66% DMSO : 33% THF (% v/v), in order to make a slurry. The startingpH of each slurry was measured using a pH meter. The pH of each vial wasadjusted to the required pH using aliquots of acetic acid or 1M sodiumhydroxide solution. If dissolution was observed at a selected pH,additional Pattern 7 solid was added to the vial, until a slurrypersisted or a maximum of 500 mg had been added. The pH was thenreadjusted as required. Recovered solids were determined to have highpurity (% area), with the majority of the experiments returning valuesof 99.5 and 99.4%. Purity values did not differ significantly between pHvalues with the exception of the solid from pH 4, where the % areapurity was determined to be 99.9%. Concentration analysis of thefiltered mother liquors showed that the concentration values generallydecreased (i.e. decreased solubility) as the pH of the system wasincreased. Sticky, highly viscous slurries were observed at pH 4 and pH8. The above technique was used to generate the diffractograms in FIG.67, FIG. 68 and FIG. 69.

Example 15. Crystallization of Set 1

Small scale crystallization trials were carried out on Pattern 7 withSet 1 to determine the effect of altering the starting concentration ofPattern 7 in DMSO/THF solvent systems. Several concentrations of Pattern7 in DMSO/THF were prepared, with starting concentrations of 100, 150and 200 mg/mL respectively. Samples were prepared in a volume of 66%DMSO: 33% THF (% v/v) and Pattern 7 (200 mg) was added as theappropriate starting concentration. Acetic acid (100 μL aliquots) wasadded at ambient temperature to each sample, until dissolution wasachieved. At each of the three concentrations, solid was crystallizedfrom solution by using a) a 1:1 equivalent ratio of acetic acid to base(1M sodium hydroxide solution), orb) adjusting the final pH to 12 using1M sodium hydroxide. The pH of the solution was measured using a pHmeter and adjusted to pH 12 using 1M sodium hydroxide solution.

The sample sets were stirred at ambient temperature for ca. 18 hourswith agitation provided via a magnetic stirrer plate. After ca. 18hours, stirring was stopped and the supernatants were filtered using0.45 μm PVDF needle filters and syringes. The concentration of thesupernatants was analyzed by HPLC. The remaining slurries in the vialswere each transferred into 0.22 μm nylon filter centrifuge tubes and thesolids were isolated by centrifugation, and the recovered solids wereanalyzed by XRPD, PLM and HPLC for purity. The above technique was usedto generate the images in FIG. 70 and FIG. 71.

TABLE 16 Set 1: pH of 12 Starting Acetic 1M NaOH Starting volumeStarting Acid solution free base (DMSO:THF Conc. added added Final mass(mg) 66/33) mL) (mg/mL) (mL) (mL) pH 200 2 100 2.4 39 12.26 200 1.3 1501.8 33.5 12.13 200 1 200 1.2 23 12.31

TABLE 17 Set 1: 1:1 acid to base ratio Starting Acetic 1M NaOH volumeStarting Acid Acetic solution (DMSO:THF Conc. added Acid added Final66/33) mL (mg/mL) (mL) Equiv. (mL) pH 2 100 2.4 94 42.1 12.42 1.3 1501.8 70.5 31.6 12.22 1 200 1.2 47 21 12.02

Example 16. Crystallization Set 2

Small scale crystallization trials were carried out on Pattern 7 withSet 2 focusing on the most suitable pH range of Pattern 7. The startingconcentration of Set 2 of crystallization experiments was lowered to 44mg/mL and the final pH was also lowered from 12 to 7, in order to reducethe volume of base required.

Samples (1 and 2) was prepared by adding Pattern 7 (200 mg) with DMSO (3mL) and THF (1.6 mL) for a slurry to be formed. The resulting slurrieswere heated to 70° C. using a temperature-controlled block. Acetic acidwas added until dissolution was achieved. The experiments were held for1 hour at 70° C. After 1 hour, Sample 1 was added 1M sodium hydroxidesolution in 100 μL aliquots, until the experiment reached pH 7. Theexperiment was then cooled from 70° C. to 25° C. at a rate of 0.25°C./min. Sample 2 solution was cooled from 70° C. to 25° C. at a rate of0.25° C./min. Once this experiment reached 25° C., 1M sodium hydroxidesolution was added in 100 μL aliquots until the experiment reached pH 7.Both samples were stirred at ambient temperature for ca. 18 hours withagitation provided via a magnetic stirrer plate. After ca. 18 hours,stirring was stopped and the supernatants were filtered using 0.45 μmPVDF needle filters and syringes. The concentration of the supernatantswas analyzed by HPLC. The remaining slurries in each sample was eachtransferred into 0.22 μm nylon filter centrifuge tubes and the solidswere isolated by centrifugation. The recovered solids were analyzed byXRPD, PLM and HPLC for purity. The above technique was used to generatethe 2^(nd) and 3^(rd) diffractograms in FIG. 72.

TABLE 18 Set 2 Starting Acetic 1M Sodium FB Volume Starting AcidHydroxide Mass (DMSO:THF Conc. Added Solution Added Final VialExperiment (mg) 66/33) (mg/mL) (mL) (mL) pH 1 1M sodium 200 4.57 44 2.85mL 52.5 7.06 hydroxide added 2 1M sodium 200 4.57 44 2.85 mL 52.5 7.38hydroxide added

Example 17. Crystallization Set 3

Small scale crystallization trials were carried out on Pattern 7 withSet 3 to determine the effect of seeding, whereby both cooling andisothermal processing was used. Samples (1 and 2) were prepared byadding Pattern 7 (200 mg) and 1 mL of 66% DMSO : 33% THF (% v/v) for aslurry to be formed. Acetic acid was added in 100 μL aliquots at ambienttemperature until dissolution was noted. Sodium hydroxide solution (1M)was added in 100 μL aliquots until the pH reached pH 5. The solution wasthen seeded with 4 mg (2%) of Pattern 7 and the pH was adjusted furtherwith 1M sodium hydroxide solution until pH 7 was achieved. Sample 1 wasstirred at ambient temperature for ca. 18 hours with agitation providedvia a magnetic stirrer plate. Sample 2 was cooled at 5° C. at 0.1°C./min and stirred for approximately 18 hours. After ca. 18 hours,stirring was stopped and the supernatants of each sample were filteredusing 0.45 μm PVDF needle filters and syringes. The concentration of thesupernatants was analyzed by HPLC. The remaining slurries from eachsample were each transferred into 0.22 μm nylon filter centrifuge tubesand the solids were isolated by centrifugation. The recovered solid wasanalyzed by XRPD, PLM and HPLC for purity. The above technique was usedto generate the 3rd and 4th diffractograms in FIG. 72.

TABLE 19 Set 3 1M Sodium 1M Sodium Hydroxide Hydroxide Acetic SolutionSolution Starting Acid Added to Added to Conc. Added reach pH 5 reach pH7 Final Vial Experiment (mg/mL) (mL) (mL) pH (mL) pH 1 Ambient 200 1.318 5.07 5.7 7.18 agitation 2 Cooling 200 1.3 18 5.09 5.5 7.04 withagitation

Example 18. Crystallization Set 4

Small-scale crystallizations were carried out on Pattern 7 with Set 4using higher concentrations (5M and 10M) of sodium hydroxide solution,in order to decrease the volume required to reach the desired pH.Samples were prepared by adding Pattern 7 (200 mg) and added 1 mL of 66%DMSO : 33% THF (% v/v) at ambient temperature for a slurry to be formed.Acetic acid was added in 100 μL aliquots at ambient temperature untildissolution was noted. Sodium hydroxide solution (5M) was then added in100 μL aliquots, until the pH of the system reached pH 5. The solutionwas then seeded with Pattern 7 (4 mg or 2%) and the pH of the sampleswas adjusted to pH 7 by the addition of sodium hydroxide solution(NaOH); whereby Vial 1 contained 5M NaOH and Vial 2 contained 10M NaOH.Samples were stirred at ambient temperature for ca. 18 hours withagitation provided via a magnetic stirrer plate. After ca. 18 hours,stirring was stopped and the formation of any solid was isolated byvacuum filtration, using a Büchner funnel and Whatman grade 1 filterpaper. The recovered solid was analyzed by XRPD, PLM and HPLC forpurity, and the concentration of the supernatants was analyzed by HPLC.The above technique was used to generate the diffractograms in FIG. 73.

TABLE 20 Set 4 Sodium Sodium Hydroxide Hydroxide Acetic SolutionSolution Starting Acid Added to Added to Conc. Added reach pH 5 reach pH7 Final Vial Experiment (mg/mL) (mL) (mL) pH (mL) pH 1 5M sodium 200 1.31.45 5 3.1 7.05 hydroxide 2 10M 200 1.3 0.51 5 1.6 7.02 sodium hydroxide

Example 19. Maturation Experimentation

100 μL aliquots of selected solvents were added to 40 mg of amorphousPattern 1 until a mobile slurry was formed. The slurries weretemperature cycled between ambient temperature and 40° C. for 72 hours.The samples were then collected, and the observed solids were isolatedand then analyzed by XRPD.

The samples were then dried and reanalyzed by XRPD. To conduct thedrying the XRPD plate was placed in an oven at 40° C. and allowed to dryfor ca. 4 hours, after which the dried samples were analyzed by RFD. Theabove technique was used to generate the diffractograms in FIG. 16, FIG.17, FIG. 18, FIG. 19, FIG. 20, FIG. 21, FIG. 22, FIG. 23, FIG. 24, FIG.25, FIG. 26, FIG. 27, FIG. 28, and FIG. 29.

TABLE 21 Observations from maturation experiments Volume of Sam- SolventInitial Observations ple Solvent System (μL) ObservationsPost-Maturation 1 1,4-Dioxane 1000 Yellow slurry Pale yellow slurry 2Acetone/water 500 Yellow slurry Yellow slurry (20:80% v/v) 3 2,2,2- 200Yellow slurry Yellow solution Trifluoroethanol 4 2-Butanone 1000 Yellowslurry Yellow slurry 5 2- 1000 Pale yellow slurry Yellow slurryEthoxyethanol 6 2-Propanol 1000 Yellow slurry Pale yellow slurry 7Acetone 1300 Yellow slurry Pale yellow slurry 8 Acetonitrile 1300 Yellowslurry Pale yellow slurry 9 Acetonitrile/ 1000 Yellow slurry Yellowslurry water (90:10% v/v) 10 Anisole 1200 Yellow slurry Yellow slurry 11Cumene 1200 Yellow slurry Yellow slurry 12 Dichloromethane 700 Yellowslurry Pale yellow slurry 13 Ethanol 700 Yellow slurry Yellow slurry 14Ethanol/water 1300 Pale yellow slurry Yellow slurry (85:15% v/v) 15Ethyl Acetate 1500 Pale yellow slurry Yellow slurry 16 Heptane 1500Yellow slurry Yellow slurry 17 Isopropyl 400 Yellow slurry Yellow slurryacetate 18 Methanol 300 Pale yellow slurry Yellow slurry 19 Methanol/400 Pale yellow slurry Yellow slurry water (95:5% v/v) 20 2N HCl 1500Pale yellow slurry Yellow slurry 21 Methyl isobutyl 1500 Yellow slurryYellow slurry ketone 22 t-Butylmethyl 1500 Yellow slurry Yellow slurryether 23 Tetrahydrofuran 1500 Yellow slurry Yellow slurry 24 Toluene1500 Yellow slurry Pale yellow slurry

TABLE 22 XRPD results after maturation experiments XRPD Post- XRPD Post-Sample Solvent System Maturation Drying 1 1,4-Dioxane Converting to 2Converting to 2 2 Acetone/water (20:80% Insufficient Insufficient v/v)solid solid 3 2,2,2-Trifluoroethanol No solid No solid 4 2-ButanonePattern 2 Pattern 2 5 2-Ethoxyethanol Pattern 2 Pattern 2 6 2-PropanolPattern 3 Pattern 3 7 Acetone Pattern 2 Pattern 2 8 Acetonitrile Pattern4 Pattern 2 9 Acetonitrile/water Pattern 1 Pattern 1 (90:10% v/v) 10Anisole Weak Pattern 5 with extra peak 11 Cumene Weak Pattern 5 withextra peak 12 Dichloromethane Pattern 2 Pattern 2 13 Ethanol Pattern 3Pattern 3 (amorphous halo) 14 Ethanol/water (85:15% Pattern 1 (HighlyPattern 1 (Highly v/v) crystalline) crystalline) 15 Ethyl AcetatePattern 2 Pattern 5 (amorphous halo) 16 Heptane Weak Pattern Pattern 3and Pattern 6 17 Isopropyl acetate Pattern 2 Pattern 2 18 MethanolPattern 1 (Highly Pattern 1 (Highly crystalline) crystalline) 19Methanol/water (95:5% Pattern 1 Pattern 1 v/v) 20 2N HCl Pattern 1Pattern 1 21 Methyl isobutyl ketone Pattern 5 Pattern 5 22 t-Butylmethylether Weak Pattern Weak Pattern 23 Tetrahydrofuran Pattern 2 Pattern 224 Toluene Weak Pattern Pattern 5

Example 20. Thermodynamic Solubility

Thermodynamic solubility was carried out for Pattern 1, Pattern 2/6mixture, Pattern 3, amorphous di-HCl salts and Pattern 7 in water at pH4.2. All samples were prepared at 30 mg/mL. Initial observations weremade. The samples were agitated at ambient temperature for ca. 24 hours.The samples were collected and observations made. Observed solids wereisolated via centrifugation. The mother liquor from the filtered samplewas analyzed by HPLC. The pH 4.3 solution was prepared by dilutingglacial acetic acid (11.6 mL) to 100 mL in deionized water, then addingsodium acetate (1.07 g) and acetic acid solution (5.9 mL) preparing upto 500 mL with deionized water; then the pH was adjusted to 4.2. Theabove technique was used to generate the images in FIG. 60.

TABLE 23 Observations and results from thermodynamic solubilityexperiments Observations at Sample Form Observations at T = 0 hours T =24 hours 1 Pattern 1 Yellow slurry Yellow solution 2 Pattern 2/6 Yellowsolution (dissolved Yellow solution upon addition) 3 Pattern 3 Yellowsolution (dissolved Yellow solution upon addition) 4 Amorphous Yellowsolution (dissolved Yellow solution Di-HCl upon addition) 5 Pattern 7Off-white slurry Off-white slurry

Example 21. Summary of Crystallization Experiments on Compound 1 FreeBase Material

The experiments from Examples 17, 18, 19, and 20 are summarized below.

TABLE 24 Summary of crystallization conditions Target pH for StartingBase precip- Set Exp. conc. used itation Experimental Summary 1 1 100 1M12 Sodium hydroxide added to pH 12 2 100 1M 12 1:1 ratio acid to base 3150 1M 12 Sodium hydroxide added to pH 12 4 150 1M 12 1:1 ratio acid tobase 5 200 1M 12 Sodium hydroxide added to pH 12 6 200 1M 12 1:1 ratioacid to base 2 7  44 mg/mL 1M 7 Experiment heated to 70° C. afterDMSO/THF addition. Base added after cooling to 25° C. 8  44 mg/mL 1M 7Experiment heated to 70° C. after DMSO/THF addition. Base added aftercooling to 25° C. 3 9 200 mg/mL 1M 7 Experiment seeded at pH 5 (2%).Ambient agitation post-seed. 10 200 mg/mL 1M 7 Experiment seeded at pH 5(2%). Cooled to 5° C. post-seed. 4 11 200 mg/mL 5M 7 Experiment seededat pH 5 (2%). Ambient agitation post-seed. 12 200 mg/mL 10M  7Experiment seeded at pH 5 (2%). Ambient agitation post-seed.

Example 22. Formation of Pattern 11

A saturated solution of Compound 1 di-HCl was prepared inN,N′-dimethylacetamide at ambient temperature and the solution was thencooled from 60° C. to 25° C. at 0.1° C. per minute. The sample was leftat ambient temperature until large crystals were obtained (roughly twoweeks). During an attempt to carry out polarized light microscopy, thecrystals re-dissolved upon agitation of the solution. The solution wasplaced in the fridge for one week to encourage crystal growth, but nocrystals appeared. Finally, ca. 10 drops of acetone were added asanti-solvent and crystals were yielded.

Example 23. Single Crystal X-Ray Analysis of Pattern 11

A suitable crystal of Pattern 11 was selected and mounted in a loopusing paratone oil. Data were collected using a Bruker D8Venturediffractometer equipped with a Photon III detector operating inshutterless mode at 100(2) K with Cu-Kα radiation (1.54178 Å). Thestructure was solved in the Olex2 software package with the She1XT(intrinsic phasing) structure solution program and refined with theShe1XL3 refinement package using Least Squares minimization. Data werecollected, solved and refined in the triclinic space-group P-1.

All non-hydrogen atoms were located in the Fourier map and theirpositions refined prior to describing the thermal movement of allnon-hydrogen atoms anisotropically. Within the structure, two completeG1T28 dihydrochloride formula units were refined. All fully occupiedhydrogen atoms were placed in calculated positions using a riding modelwith fixed Uiso at 1.2 times for all CH, CH₂ and NH groups and 1.5 timesfor all CH₃ groups. All partially occupied hydroxyl hydrogen atoms werelocated in the Fourier map and their positions and occupancy refined.

The highest residual Fourier peak was found to be 0.54 e.Å⁻³ approx.1.19 Å from C(1) and the deepest Fourier hole was found to be −0.29e.Å⁻³ approx. 0.63 Å from C1(2).

Crystal Data

C₂₄H₃₃C₁₂N₈O_(1.5) (M=528.48 g/mol): triclinic, space group P-1 (no. 2),a=12.2824(2) Å, b=14.5593(3) Å, c=15.9459(3) Å, α=72.0580(10)°,β=85.7150(10)°, γ=74.6560(10)°, V=2616.05(9) Å3, Z=4, T=100.0 K,μt(CuKα)=2.523 mm-1, Dcalc=1.342 g/cm³, 77718 reflections measured(6.598°≤2Θ≤149.518°), 10714 unique (R_(int)=0.0685, R_(sigma)=0.0357)which were used in all calculations. The final R₁ was 0.0461 (I>2σ(I))and wR₂ was 0.1363 (all data).

TABLE 25 Crystallographic parameters to refine Pattern 11 Pattern 11(di-HCl semihydrate) Empirical formula C₂₄H₃₃Cl₂N₈O_(1.5) Formula weight528.48 Temperature/K 100 Crystal system triclinic Space group P-1 a/Å12.2824(2) b/Å 14.5593(3) c/Å 15.9459(3) α/° 72.0580(10) β/° 85.7150(10)γ/° 74.6560(10) Volume/Å³ 2616.05(9) Z 4 ρcalcg/cm³ 1.342 μ/mm⁻¹ 2.523F(000) 1116 Crystal size/mm³ 0.18 × 0.08 × 0.02 Radiation CuKα (λ =1.54178) 2Θ range for data collection/° 6.598 to 149.518 Index ranges−15 ≤ h ≤ 15, −18 ≤ k ≤ 18, −19 ≤ l ≤ 19 Reflections collected 77718Independent reflections 10714 [R_(int) = 0.0685, R_(sigma) = 0.0357]Data/restraints/parameters 10714/0/633 Goodness-of-fit on F² 1.038 FinalR indexes [I >= 2σ (I)] R1 = 0.0461, wR2 = 0.1260 Final R indexes [alldata] R1 = 0.0613, wR2 = 0.1363 Largest diff. peak/hole/e Å⁻³ 0.54/−0.29

Example 24. Single Crystal Determination Results for Pattern 11

The hemihydrate of Compound 1 as a dihydrochloride was found to beextremely crystalline and possess unit cell dimensions as follows:

Triclinic P-1 a = 12.2824(2) A a = 72.0580(10)° b = 14.5593(3) A b =85.7150(10)° c = 15.9459(3) A a = 74.6560(10)° Volume = 2,616.05 (9) A³Z = 4, Z′ = 2

This asymmetric unit contains two complete Compound 1 formula units andtwo chloride anions. The solvent mask was calculated and 91.1 electronswere found within a volume of 341.0 Å₃ in a void in the unit cell. Thiscorresponds to 45.5 electrons per asymmetric unit and 22.75 electronsper G1T28 formula unit. This is consistent with the presence of oneadditional chloride anion and a water molecule at 50% occupancy perG1T28 molecule, totaling 44 electrons per asymmetric unit. The solventmask was removed in order to determine the location of the disorderedchlorides which were found within channels intersecting the unit cell atthe cell origins.

The final refinement parameters were as follows:

-   -   R₁[I>2σ(I)]=4.61%    -   GooF (Goodness of fit)=1.038    -   wR₂ (all data)=13.63%    -   R_(int)=6.85%

Packing within the structure was shown to be efficient whereby amoderate density of 1.342 g/cm³ was noted. A solvent mask was applied toa channel along the a-axis which contained the disordered solvent andcounterion.

A discrete hydrogen bonding motif was noted in the asymmetric unit, asshown in FIG. 90. The hydrogen bonding association between theprotonated piperazine (H(29) or H29)′) and chloride counterion (Cl(1) orCl(2)), and between the amide hydrogen (H(3) or H(3)′) and chloridecounterion (Cl(1) or Cl(2)), with H(29) . . . Cl(1)/H(29)′ . . . Cl(2)and Cl(1) . . . H(3)′/Cl(2) . . . H(3) measuring 2.08(3)/2.10(3) and2.40(3)/2.36(3) Å, respectively.

In addition to the discrete cage-like hydrogen bonding associationnoted, the pyrimidine and bridging amines appeared to show a clearassociation. This interaction was found to be of moderate strength,measuring 2.15(2) A between N(9) and H(19), forming a R22(8) motif.There was no evidence of π-π interactions, the primary drive tocrystallization was via hydrogen bond formation, as described above. Theremaining interactions between molecules in the structure were made upof van de Waal's interactions. A simulated XRPD diffractogram of hasbeen calculated and compared to an experimental diffractogram of Pattern1 (FIG. 96). The diffractograms are not consistent with each other.

TABLE 26 XRPD simulated peak list for Pattern 11 No. Pos. [°2θ]d-spacing [Å] Height [cts] Rel Int. [%] 1 5.8209 15.17080 2075.83 20.532 6.5936 13.39452 10112.42 100 3 7.3734 11.97972 7190.89 71.11 4 9.25389.54913 52.20 0.52 5 10.0260 8.81529 3409.28 33.71 6 11.1511 7.92828189.13 1.87 7 11.6573 7.58514 3732.39 36.91 8 12.7374 6.94428 1333.2413.18 9 13.9012 6.36539 99.45 0.98 10 14.7220 6.01229 1164.27 11.51 1115.0305 5.88958 2108.97 20.86 12 15.1432 5.84602 1602.37 15.85 1315.9841 5.54030 3119.67 30.85 14 16.5233 5.36071 1270.96 12.57 1516.7700 5.28239 445.67 4.41 16 17.5939 5.03682 326.19 3.23 17 17.83784.96850 2822.86 27.91 18 18.5352 4.78310 994.29 9.83 19 18.9219 4.686231632.15 16.14 20 19.0676 4.65074 3212.03 31.76 21 19.4254 4.56589 860.648.51 22 19.9606 4.44465 2694.80 26.65 23 20.5317 4.32229 179.94 1.78 2420.8288 4.26130 750.55 7.42 25 21.4583 4.13769 905.33 8.95 26 21.64524.10237 471.94 4.67 27 22.0978 4.01937 297.14 2.94 28 22.4959 3.949136503.85 64.32 29 23.1964 3.83144 1588.05 15.70 30 23.9311 3.71545 236.962.34 31 24.1998 3.67479 1260.44 12.46 32 24.3822 3.64771 524.70 5.19 3325.0249 3.55548 711.00 7.03 34 25.6050 3.47621 114.77 1.13 35 26.25073.39216 712.98 7.05 36 26.5654 3.35268 1069.11 10.57 37 27.1874 3.27737990.66 9.80 38 27.3587 3.25725 428.86 4.24 39 28.0072 3.18327 272.762.70 40 28.3983 3.14033 132.18 1.31 41 28.6382 3.11455 569.96 5.64 4228.7733 3.10024 600.54 5.94 43 29.3735 3.03825 603.26 5.97 44 30.15752.96103 250.17 2.47 45 30.6752 2.91221 206.57 2.04 46 30.8126 2.89955297.80 2.94 47 31.0668 2.87640 306.34 3.03 48 31.5495 2.83348 181.121.79 49 31.7273 2.81801 153.60 1.52 50 32.3419 2.76584 316.57 3.13 5132.6990 2.73645 406.20 4.02 52 32.9597 2.71539 220.82 2.18 53 33.25442.69201 443.02 4.38 54 33.5207 2.67123 254.44 2.52 55 34.0803 2.62863397.79 3.93 56 34.2787 2.61387 121.68 1.20 57 34.6959 2.58339 141.941.40 58 34.9584 2.56459 81.57 0.81

TABLE 27 Highest intensity simulated XRPD diffractogram peaks forPattern 11 No. Pos. [°2θ] d-spacing [Å] Height Rel. Int. [%] 1 5.820915.17080 2075.83 20.53 2 6.5936 13.39452 10112.42 100 3 7.3734 11.979727190.89 71.11 4 10.0260 8.81529 3409.28 33.71 5 11.6573 7.58514 3732.3936.91 6 12.7374 6.94428 1333.24 13.18 7 14.7220 6.01229 1164.27 11.51 815.0305 5.88958 2108.97 20.86 9 15.1432 5.84602 1602.37 15.85 10 15.98415.54030 3119.67 30.85 11 16.5233 5.36071 1270.96 12.57 12 17.83784.96850 2822.86 27.91 13 18.5352 4.78310 994.29 9.83 14 18.9219 4.686231632.15 16.14 15 19.0676 4.65074 3212.03 31.76 16 19.9606 4.444652694.80 26.65 17 22.4959 3.94913 6503.85 64.32 18 23.1964 3.831441588.05 15.70 19 24.1998 3.67479 1260.44 12.46 20 26.5654 3.352681069.11 10.57

Example 24. Single Crystal Solution of Pattern 11

The coordinates and solved parameters of the single crystal solution forPattern 11 are provided below:

TABLE 28 Atom coordinates for Pattern 11 Atom x y z U(eq) Cl1 5968.1(4)7961.9(3) 4715.8(3) 27.07(11) C1 1379.6(14) 3923.2(13) 3199.1(11)19.8(3) C2 1218.2(16) 3038.6(14) 3983.3(12) 26.1(4) N3 774.0(15)2331.4(13) 3714.7(12) 31.5(4) O4 971.0(13) 1287.2(11) 2877.5(10) 34.9(3)C4 1166.4(16) 2015.1(14) 3018.8(13) 27.5(4) C5 1874.1(16) 2601.6(14)2413.9(13) 26.2(4) C6 2426.2(17) 2431.5(14) 1680.0(13) 26.9(4) C72955.6(15) 3223.7(13) 1311.4(12) 22.5(4) C8 3664.2(15) 3494.8(13)608.8(12) 23.1(4) N9 4016.6(12) 4321.3(11) 440.4(10) 21.1(3) N294003.8(12) 9560.7(12) 3720.1(10) 21.3(3) C10 3637.3(14) 4904.4(13)979.8(11) 18.7(3) N11 2977.4(12) 4724.2(11) 1682.7(10) 20.4(3) C122670.0(14) 3867.6(13) 1847.7(12) 20.4(3) N13 2042.9(13) 3467.9(11)2543.4(10) 22.4(3) C14 2044.9(15) 4496.8(14) 3537.7(12) 24.5(4) C151312.3(17) 5098.3(15) 4115.2(13) 28.9(4) C16 251.5(18) 5779.1(15)3624.6(14) 34.5(5) C17 −434.0(17) 5198.0(17) 3339.0(14) 34.7(5) C18273.0(16) 4603.2(15) 2759.6(13) 28.2(4) N19 4016.4(13) 5755.4(11)750.7(10) 21.2(3) C20 3786.8(15) 6505.3(13) 1157.8(11) 19.8(3) N214545.5(12) 7047.4(11) 996.8(10) 20.1(3) C22 4444.0(15) 7740.1(12)1413.9(11) 19.8(3) C23 3596.1(14) 7936.8(12) 2007.6(12) 19.6(3) C242757.8(15) 7413.6(14) 2111.0(13) 24.8(4) C25 2842.4(15) 6706.2(14)1684.8(13) 24.1(4) N26 3518.6(12) 8641.4(11) 2473.3(10) 20.6(3) C274464.3(17) 9099.2(15) 2348.5(13) 27.6(4) C28 4219.2(19) 9942.0(15)2762.0(13) 31.3(4) C29 3811.0(17) 10355.8(15 4159.4(15) 30.6(4) C303039.5(16) 9086.1(15) 3850.9(14) 28.1(4) C31 3300.8(16) 8258.1(14)3421.1(12) 24.6(4) C12 −741.1(4) 1809.4(4) 5414.3(3) 27.98(12) N291143.4(13) 213.1(12) 6499.5(11) 24.6(3) C1′ 3792.0(14) 5843.2(13)6922.0(11) 19.3(3) C2′ 4910.9(15) 6136.6(14) 6681.8(13) 24.4(4) N3′4798.2(14) 7190.1(13) 6583.2(11) 27.5(3) O4′ 4185.4(12) 8516.1(10)7119.1(10) 31.8(3) C4′ 4219.5(15 7648.1(14 7149.6(13) 24.5(4) C5′3557.2(15 7054.1(13 7803.5(12) 22.6(4) C6′ 2966.3(16 7280.4(148502.0(12) 24.5(4) C7′ 2371.0(15 6530.8(13 8862.6(12) 21.6(4) C8′1584.2(16 6339.9(14 9522.5(12) 23.2(4) N9′ 1139.3(13 5562.6(119667.3(10) 22.0(3) C10′ 1494.7(14 4960.0(13 9141.7(11) 18.9(3) N11′2217.1(12 5071.8(11 8477.9(10) 19.3(3) C12′ 2644.0(14 5861.3(138350.9(11) 19.2(3) N13′ 3390.2(12 6168.8(11 7711.9(10) 19.8(3) C14′2901.2(15 6362.3(13 6191.8(12) 22.5(4) C15′ 3229.4(16 6008.9(145374.4(12) 26.4(4) C16′ 3477.5(16 4872.2(14 5606.1(13) 27.0(4) C17′4368.2(16 4352.5(14 6324.6(12) 25.1(4) C18′ 4018.2(15 4703.1(137141.7(12) 23.5(4) N19′ 1005.7(13 4170.4(11 9346.8(10) 21.2(3) C20′1207.6(14 3410.0(13 8956.3(11) 19.2(3) N21′ 342.1(12) 3005.5(118986.5(10) 21.6(3) C22′ 455.3(15) 2284.2(13 8601.9(12) 21.2(3) C23′1418.4(14 1932.0(13 8165.0(11) 19.8(3) C24′ 2339.9(15 2319.0(138190.6(13) 23.3(4) C25′ 2250.1(14 3050.6(13 8589.9(12) 22.2(4) N26′1524.6(12 1195.2(11 7724.3(10) 20.4(3) C27′ 1809.5(16 1545.0(146785.6(13) 25.9(4) C28′ 2109.2(16 684.7(15) 6391.2(14) 27.4(4) C29′1380.1(18 −613.6(16) 6098.7(16) 34.2(5) C30′ 843.8(17) −116.7(14)7447.6(13) 27.6(4) C31′ 562.4(15) 759.4(13) 7821.8(12) 22.9(4)

TABLE 29 Anisotropic displacement parameters (A² × 10³) for Pattern 11.The anisotropic displacement factor exponent takes the form:−2π²[h²a*²U₁₁ + 2hka*b*U₁₂ + . . .] Atom U11 U22 U33 U23 U13 U12 Cl125.4(2) 33.0(2) 23.3(2) −7.14(18) 3.64(1  — C1 19.5(8) 25.8(9) 18.1(8)−10.7(7) 6.5(6) −9.4(7) C2 27.7(9) 30.7(10) 22.5(9) −8.0(8) 4.1(7)−12.7(8) N3 39.1(9) 33.2(9) 30.5(9) −12.5(7) 15.3(8  −23.6(8) O4 47.7(8)31.1(7) 35.6(8) −13.6(6) 13.9(7  −25.9(7) C4 31.5(9) 25.7(9) 29.3(10)−8.5(8) 6.8(8) −15.9(8) C5 30.5(9) 24.1(9) 29.0(10) −11.1(8) 8.6(8)−14.2(8) C6 34.9(10 22.6(9) 28.8(10) −12.0(8) 9.5(8) −14.1(8) C7 27.4(9)21.4(8) 22.6(9) −10.7(7) 5.6(7) −9.7(7) C8 29.8(9) 22.5(8) 22.0(9)−12.2(7) 6.5(7) −10.3(7) N9 25.9(7) 23.0(7) 19.7(7) −10.7(6) 6.3(6)−11.4(6) N29 19.9(7) 26.2(8) 23.9(8) −14.1(6) 3.9(6) −9.1(6) C10 21.3(8)20.2(8) 17.7(8) −7.7(7) 3.2(6) −9.1(6) N11 23.5(7) 24.1(7) 18.8(7)−10.1(6) 6.7(6) −12.2(6) C12 21.6(8) 22.8(8) 19.8(8) −7.9(7) 5.3(7)−10.0(7) N13 26.1(7) 23.7(7) 23.2(8) −11.7(6) 9.5(6) −13.1(6) C1423.0(8) 32.9(10) 21.9(9) −10.7(8) 4.2(7) −12.7(7) C15 34.5(10 34.7(10)23.6(9) −14.1(8) 8.0(8) −14.8(8) C16 41.6(11 30.6(10) 28.1(10) −9.8(8)10.7(9  −5.4(9) C17 25.1(9) 41.3(12) 31.9(11) −11.0(9) 0.8(8) 1.2(8) C1823.7(9) 35.8(10) 23.6(9) −8.4(8) 0.5(7) −6.1(8) N19 25.9(7) 22.4(7)20.9(8) −11.2(6) 8.8(6) −12.8(6) C20 23.2(8) 19.7(8) 18.4(8) −6.6(7)1.1(7) −7.9(6) N21 25.4(7) 20.8(7) 17.2(7) −7.2(6) 4.0(6) −10.4(6) C2223.8(8) 19.0(8) 19.5(8) −5.8(7) 2.3(7) −10.9(7) C23 21.3(8) 16.4(8)23.1(9) −7.6(7) −1.6(7)  −5.7(6) C24 18.4(8) 27.9(9) 33.6(10) −16.7(8)6.5(7) −8.0(7) C25 19.5(8) 25.9(9) 33.5(10) −15.9(8) 4.7(7) −10.0(7) N2622.2(7) 21.3(7) 23.0(8) −11.0(6) 3.3(6) −9.1(6) C27 38.0(10 34.2(10)23.2(9) −16.2(8) 13.2(8  −24.8(9) C28 47.0(12 30.7(10) 25.4(10) −11.4(8)6.1(9) −23.3(9) C29 28.6(9) 33.6(10) 40.5(12) −25.3(9) 3.8(8) −10.2(8)C30 23.2(9) 37.1(10) 36.1(11) −23.9(9) 11.8(8  −15.7(8) C31 28.1(9)27.4(9) 26.2(10) −15.4(8) 12.2(7  −15.0(7) C12 22.4(2) 37.4(2) 24.8(2)−8.01(18) 4.76(1  − N29′ 21.3(7) 30.2(8) 29.1(9) −18.4(7) 4.1(6) −8.0(6)C1′ 19.7(8) 23.0(8) 17.2(8) −8.3(7) 5.1(6) −7.5(6) C2′ 19.5(8) 30.7(9)26.3(9) −11.8(8) 5.0(7) −9.2(7) N3′ 29.1(8) 32.1(9) 26.8(8) −9.8(7)9.6(7) −18.6(7) O4′ 35.6(7) 28.4(7) 36.3(8) −10.6(6) 8.4(6) −17.8(6) C4′24.0(8) 26.7(9) 26.0(9) −7.5(7) 2.1(7) −12.6(7) C5′ 23.8(8) 24.0(9)24.3(9) −9.5(7) 3.8(7) −11.7(7) C6′ 29.4(9) 25.7(9) 24.5(9) −11.0(7)5.3(7) −14.4(7) C7′ 25.7(8) 23.0(8) 20.9(9) −10.8(7) 3.5(7) −10.4(7) C8′29.4(9) 25.6(9) 22.2(9) −14.2(7) 7.0(7) −13.1(7) N9′ 26.2(7) 24.2(7)20.9(8) −11.2(6) 7.5(6) −12.0(6) C10′ 20.4(8) 20.5(8) 18.1(8) −8.0(7)2.0(6) −7.2(6) N11′ 20.0(7) 23.2(7) 18.5(7) −9.3(6) 3.5(6) −9.0(6) C12′17.8(7) 23.5(8) 18.6(8) −8.1(7) 2.8(6) −7.9(6) N13′ 21.0(7) 22.8(7)19.6(7) −9.6(6) 5.3(6) −9.7(6) C14′ 21.7(8) 24.7(9) 23.1(9) −9.5(7)2.6(7) −7.2(7) C15′ 30.3(9) 28.7(9) 20.1(9) −6.9(7) 1.2(7) −8.1(8) C16′29.4(9) 30.1(10) 25.7(10) −13.5(8) 8.3(8) −10.8(8) C17′ 28.1(9) 23.0(9)25.5(9) −10.2(7) 11.0(7  −8.0(7) C18′ 25.7(9) 20.6(8) 22.4(9) −6.3(7)5.9(7) −4.7(7) N19′ 23.2(7) 24.9(7) 22.2(8) −13.0(6) 9.7(6) −12.8(6)C20′ 22.4(8) 20.4(8) 17.1(8) −6.8(6) 3.3(6) −8.9(6) N21′ 24.6(7) 24.1(7)21.1(7) −10.9(6) 8.0(6) −11.9(6) C22′ 24.7(8) 23.8(8) 20.7(9) −9.5(7)5.6(7) −13.3(7) C23′ 23.2(8) 19.2(8) 19.3(8) −7.6(7) 2.0(7) −7.7(7) C24′18.5(8) 24.4(9) 29.3(10) −13.0(7) 4.3(7) −4.4(7) C25′ 19.2(8) 23.9(9)28.2(9) −12.1(7) 1.9(7) −8.7(7) N26′ 21.8(7) 22.6(7) 21.7(8) −11.9(6)3.8(6) −8.7(6) C27′ 29.5(9) 28.7(9) 26.4(10) −15.3(8) 11.6(7  −14.0(8)C28′ 24.4(9) 34.7(10) 32.7(10) −20.9(9) 9.1(8) −13.1(8) C29′ 31.5(1039.1(11) 47.3(13) −32.2(10) 6.9(9) −13.6(9) C30′ 32.9(10 25.5(9)30.7(10) −13.7(8) 5.3(8) −13.4(8) C31′ 26.5(9) 24.7(9) 24.1(9) −12.4(7)5.8(7) −13.1(7) N29′ 21.3(7) 30.2(8) 29.1(9) −18.4(7) 4.1(6) −8.0(6) C1′19.7(8) 23.0(8) 17.2(8) −8.3(7) 5.1(6) −7.5(6) C2′ 19.5(8) 30.7(9)26.3(9) −11.8(8) 5.0(7) −9.2(7) N3′ 29.1(8) 32.1(9) 26.8(8) −9.8(7)9.6(7) −18.6(7) O4′ 35.6(7) 28.4(7) 36.3(8) −10.6(6) 8.4(6) −17.8(6) C4′24.0(8) 26.7(9) 26.0(9) −7.5(7) 2.1(7) −12.6(7) C5′ 23.8(8) 24.0(9)24.3(9) −9.5(7) 3.8(7) −11.7(7) C6′ 29.4(9) 25.7(9) 24.5(9) −11.0(7)5.3(7) −14.4(7) C7′ 25.7(8) 23.0(8) 20.9(9) −10.8(7) 3.5(7) −10.4(7) C8′29.4(9) 25.6(9) 22.2(9) −14.2(7) 7.0(7) −13.1(7) N9′ 26.2(7) 24.2(7)20.9(8) −11.2(6) 7.5(6) −12.0(6) C10′ 20.4(8) 20.5(8) 18.1(8) −8.0(7)2.0(6) −7.2(6) N11′ 20.0(7) 23.2(7) 18.5(7) −9.3(6) 3.5(6) −9.0(6) C12′17.8(7) 23.5(8) 18.6(8) −8.1(7) 2.8(6) −7.9(6) N13′ 21.0(7) 22.8(7)19.6(7) −9.6(6) 5.3(6) −9.7(6) C14′ 21.7(8) 24.7(9) 23.1(9) −9.5(7)2.6(7) −7.2(7) C15′ 30.3(9) 28.7(9) 20.1(9) −6.9(7) 1.2(7) −8.1(8) C16′29.4(9) 30.1(10) 25.7(10) −13.5(8) 8.3(8) −10.8(8) C17′ 28.1(9) 23.0(9)25.5(9) −10.2(7) 11.0(7  −8.0(7) C18′ 25.7(9) 20.6(8) 22.4(9) −6.3(7)5.9(7) −4.7(7) N19′ 23.2(7) 24.9(7) 22.2(8) −13.0(6) 9.7(6) −12.8(6)C20′ 22.4(8) 20.4(8) 17.1(8) −6.8(6) 3.3(6) −8.9(6) N21′ 24.6(7) 24.1(7)21.1(7) −10.9(6) 8.0(6) −11.9(6) C22′ 24.7(8) 23.8(8) 20.7(9) −9.5(7)5.6(7) −13.3(7) C23′ 23.2(8) 19.2(8) 19.3(8) −7.6(7) 2.0(7) −7.7(7) C24′18.5(8) 24.4(9) 29.3(10) −13.0(7) 4.3(7) −4.4(7) C25′ 19.2(8) 23.9(9)28.2(9) −12.1(7) 1.9(7) −8.7(7) N26′ 21.8(7) 22.6(7) 21.7(8) −11.9(6)3.8(6) −8.7(6) C27′ 29.5(9) 28.7(9) 26.4(10) −15.3(8) 11.6(7  −14.0(8)C28′ 24.4(9) 34.7(10) 32.7(10) −20.9(9) 9.1(8) −13.1(8) C29′ 31.5(1039.1(11) 47.3(13) −32.2(10) 6.9(9) −13.6(9) C30′ 32.9(10 25.5(9)30.7(10) −13.7(8) 5.3(8) −13.4(8) C31′ 26.5(9) 24.7(9) 24.1(9) −12.4(7) 58(7) −13.1(7)

TABLE 30 Bond lengths for Pattern 11 Atom Atom Length/Å C1 C2 1.539(2)C1 N13 1.487(2) C1 C14 1.537(2) C1 C18 1.524(3) C2 N3 1.465(2) N3 C41.339(3) O4 C4 1.233(2) C4 C5 1.475(2) C5 C6 1.364(3) C5 N13 1.408(2) C6C7 1.421(2) C7 C8 1.390(2) C7 C12 1.417(2) C8 N9 1.329(2) N9 C101.363(2) N29 C28 1.486(2) N29 C29 1.489(2) N29 C30 1.494(2) C10 N111.333(2) C10 N19 1.373(2) N11 C12 1.341(2) C12 N13 1.369(2) C14 C151.543(2) C15 C16 1.511(3) C16 C17 1.518(3) C17 C18 1.527(3) N19 C201.393(2) C20 N21 1.336(2) C20 C25 1.401(2) N21 C22 1.345(2) C22 C231.387(2) C23 C24 1.407(2) C23 N26 1.421(2) C24 C25 1.377(2) N26 C271.458(2) N26 C31 1.472(2) C27 C28 1.518(3) C30 C31 1.516(2) N29′ C28′1.495(2) N29′ C29′ 1.484(2) N29′ C30′ 1.488(2) C1′ C2′ 1.533(2) C1′ N13′1.485(2) C1′ C14′ 1.528(2) C1′ C18′ 1.539(2) C2′ N3′ 1.463(2) N3′ C4′1.338(2) O4′ C4′ 1.239(2) C4′ C5′ 1.479(2) C5′ C6′ 1.362(3) C5′ N13′1.409(2) C6′ C7′ 1.424(2) C7′ C8′ 1.389(2) C7′ C12′ 1.416(2) C8′ N9′1.334(2) N9′ C10′ 1.363(2) C10′ N11′ 1.328(2) C10′ N19′ 1.376(2) N11′C12′ 1.340(2) C12′ N13′ 1.369(2) C14′ C15′ 1.533(2) C15′ C16′ 1.530(3)C16′ C17′ 1.516(3) C17′ C18′ 1.534(2) N19′ C20′ 1.390(2) C20′ N21′1.336(2) C20′ C25′ 1.406(2) N21′ C22′ 1.344(2) C22′ C23′ 1.387(2) C23′C24′ 1.399(2) C23′ N26′ 1.428(2) C24′ C25′ 1.377(2) N26′ C27′ 1.473(2)N26′ C31′ 1.460(2) C27′ C28′ 1.519(2) C30′ C31′ 1.519(2)

TABLE 31 Bond angles for Pattern 11 Atom Atom Atom Angle/° N13 C1 C2105.39(14) N13 C1 C14 111.19(13) N13 C1 C18 107.77(14) C14 C1 C2107.44(15) C18 C1 C2 113.32(15) C18 C1 C14 111.60(15) N3 C2 C1112.63(15) C4 N3 C2 122.86(16) N3 C4 C5 115.08(16) O4 C4 N3 123.95(17)O4 C4 C5 120.96(18) C6 C5 C4 128.19(17) C6 C5 N13 110.69(15) N13 C5 C4121.11(16) C5 C6 C7 106.47(16) C8 C7 C6 137.25(17) C8 C7 C12 115.52(15)C12 C7 C6 107.21(15) N9 C8 C7 121.79(16) C8 N9 C10 117.04(15) C28 N2 C29111.98(15) C28 N2 C30 109.42(15) C29 N2 C30 112.00(14) N9 C1 N19113.54(15) N11 C1 N9 127.31(15) N11 C1 N19 119.14(15) C10 N1 C12113.82(14) N11 C1 C7 124.37(16) N11 C1 N13 126.83(16) N13 C1 C7108.80(14) C5 N1 C1 121.69(14) C12 N1 C1 129.83(14) C12 N1 C5 106.71(14)C1 C1 C15 111.95(14) C29′ N29′ C28′ 111.90(14) C29′ N29′ C30′ 112.09(16)C30′ N29′ C28′ 109.99(15) C2′ C1′ C18′ 108.89(14) N13′ C1′ C2′104.98(13) N13′ C1′ C14′ 109.06(14) N13′ C1′ C18′ 110.78(14) C14′ C1′C2′ 112.98(14) C14′ C1′ C18′ 110.05(14) N3′ C2′ C1′ 112.38(15) C4′ N3′C2′ 122.79(16) N3′ C4′ C5′ 114.67(16) O4′ C4′ N3′ 123.90(17) O4′ C4′ C5′121.36(17) C6′ C5′ C4′ 128.25(17) C6′ C5′ N13′ 110.60(15) N13′ C5′ C4′120.95(16) C5′ C6′ C7′ 106.45(16) C8′ C7′ C6′ 136.96(17) C8′ C7′ C12′115.55(15) C12′ C7′ C6′ 107.36(15) N9′ C8′ C7′ 121.54(16) C8′ N9′ C10′117.10(15) N9′ C10′ N19′ 113.09(15) N11′ C10′ N9′ 127.35(15) N11′ C10′N19′ 119.56(15) C10′ N11′ C12′ 113.82(14) N11′ C12′ C7′ 124.61(15) N11′C12′ N13′ 126.81(15) N13′ C12′ C7′ 108.58(14) C5′ N13′ C1′ 121.28(14)C12′ N13′ C1′ 130.41(14) C12′ N13′ C5′ 106.96(14) C1′ C14′ C15′111.80(15)

TABLE 32 Bond angles for Pattern 11 continued Atom Atom Atom Angle/° C16C15 C14 110.47(16) C15 C16 C17 111.59(17) C16 C17 C18 110.50(17) C1 C18C17 112.70(16) C10 N19 C20 128.14(15) N19 C20 C25 124.39(15) N21 C20 N19114.26(15) N21 C20 C25 121.34(16) C20 N21 C22 118.67(15) N21 C22 C23124.36(15) C22 C23 C24 115.77(15) C22 C23 N26 124.13(15) C24 C23 N26120.06(15) C25 C24 C23 120.58(16) C24 C25 C20 118.84(16) C23 N26 C27115.10(14) C23 N26 C31 113.71(14) C27 N26 C31 109.67(14) N26 C27 C28110.71(15) N29 C28 C27 110.27(15) N29 C30 C31 109.70(14) N26 C31 C30111.14(15) C16′ C15′ C14′ 111.55(15) C17′ C16′ C15′ 111.64(16) C16′ C17′C18′ 110.55(15) C17′ C18′ C1′ 111.87(15) C10′ N19′ C20′ 127.48(15) N19′C20′ C25′ 123.57(15) N21′ C20′ N19′ 114.77(15) N21′ C20′ C25′ 121.60(16)C20′ N21′ C22′ 118.62(15) N21′ C22′ C23′ 124.05(15) C22′ C23′ C24′116.36(15) C22′ C23′ N26′ 124.19(15) C24′ C23′ N26′ 119.43(15) C25′ C24′C23′ 120.60(16) C24′ C25′ C20′ 118.47(16) C23′ N26′ C27′ 112.98(14) C23′N26′ C31′ 114.98(14) C31′ N26′ C27′ 109.96(14) N26′ C27′ C28′ 110.61(15)N29′ C28′ C27′ 109.76(14) N29′ C30′ C31′ 110.37(15) N26′ C31′ C30′110.07(14)

TABLE 33 Hydrogen positions for Pattern 11 Ato x y z U(eq H2 1952.822684.00 4287.21 31 H2  692.24 3296.46 4408.15 31 H3  290(20)  2055(18) 4088(17) 38 H6 2451.78 1890.37 1458.87 32 H8 3903.14 3078.05  238.95 28H29 4630(19  9044(17)  3990(15) 26 H14 2330.99 4958.46 3028.79 29 H142703.91 4017.82 3886.35 29 H15 1107.50 4633.61 4664.44 35 H15 1748.765501.07 4278.38 35 H16  458.50 6273.83 3098.44 41 H16 −214.76 6147.554009.52 41 H17 — 5666.28 3006.91 42 H17 −689.43 4736.56 3865.85 42 H18−171.39 4192.92 2617.73 34 H18  440.12 5073.20 2198.81 34 H19 4530(20 5778(16)  399(15) 25 H22 4989.26 8119.89 1293.93 24 H24 2127.98 7549.402477.86 30 H25 2270.39 6360.61 1747.02 29 H27 4603.62 9362.95 1710.35 33H27 5153.40 8588.81 2620.46 33 H28 4870.71 10241.40  2673.81 38 H283550.90 10467.89  2471.64 38 H29 3108.17 10859.93  3935.86 46 H294443.11 10670.00  4033.98 46 H29 3754.81 10060.90  4797.45 46 H302344.25 9593.22 3586.92 34 H30 2910.10 8809.41 4488.97 34 H31 3971.227734.39 3710.35 30 H31 2655.36 7951.83 3502.74 30 H29  560(20)  692(18) 6199(16) 29 H2′ 5477.81 5719.97 7147.31 29 H2′ 5189.48 5999.15 6122.6929 H3′ 5130(20 7539(18  6144(16) 33 H6′ 2953.53 7829.77 8708.09 29 H8′1357.40 6772.85 9879.05 28 H14 2805.30 7092.82 6025.24 27 H14 2169.106223.95 6415.63 27 H15 2606.17 6321.08 4936.54 32 H15 3906.18 6227.175103.58 32 H16 2773.98 4662.18 5805.18 32 H16 3740.12 4667.85 5072.76 32H17 5097.07 4500.45 6103.61 30 H17 4471.84 3621.32 6485.66 30 H184624.61 4376.30 7592.81 28 H18 3327.54 4497.13 7393.07 28 H19  410(20)4234(16  9628(15) 25 H22 −162.62 1997.61 8631.02 25 H24 3033.65 2074.817930.26 28 H25 2878.04 3306.40 8617.03 27 H27 2456.74 1846.14 6723.87 31H27 1158.30 2064.81 6461.48 31 H28 2280.62  933.41 5757.37 33 H282788.87  182.57 6690.01 33 H29 2084.80 — 6345.00 51 H29 1453.35 −345.665459.06 51 H29  758.50 −940.55 6227.50 51 H30 1485.34 −641.08 7780.47 33H30  185.78 −404.85 7512.32 33 H31 −101.53 1270.66 7507.69 27 H31 370.70  530.13 8453.40 27

Example 25. Single Crystal X-Ray Crystallization Experiment ConditionsUsed on Pattern 1

Diffraction data were collected on XRD1 beamline (Lausi, et al, 2015),in its standard single crystal configuration. The experimental setupconsists in a Huber goniometer with κ geometry, fully controllable fromremote. Samples diffraction properties were characterized at 100K(nitrogen stream supplied through an Oxford Cryostream 700—OxfordCryosystems Ltd., Oxford, United Kingdom) through the rotating crystalmethod.

Data were acquired using a monochromatic wavelength of 0.6199 Å, on aPilatus 6M hybrid-pixel area detector (DECTRIS Ltd., Baden-Daettwil,Switzerland). Crystals were dipped in NHV oil (Jena Bioscience, Jena,Germany), mounted at RT on kapton loops (MiTeGen, Ithaca, USA) and flashfrozen in liquid nitrogen. Diffraction properties of 15 differentcrystals have been assessed—all of them appear as thin yellow needlesless than 50-80 mm in the longest direction. Most of specimensdiffracted around ˜0.9-1 Å, but few of them diffracted up to ˜0.7-0.8 Å.Standard “Omega-scan” XRD data have been collected on the bestdiffracting ones, till radiation damage started degrading the dataquality significantly. An oscillation range of 0.5° has been chosen foroptimal Bragg peaks separation. The same crystalline form has been foundin crystals from all the three batches supplied.

The structure was solved using direct methods by SHELXT-2014/7(Sheldrick, G. M., 2015a) and refined by least-square full matrixrefinement using SHELXL-2014/7 (Sheldrick, G. M., 2015b). All H-atoms,except for those linked to N or O atoms, were included from thegeometry. In the final cycles of refinement, the large peak ˜2 e/Å3 wasfound nearby ClB atom. This was interpreted as partial disorder of thisatom. The refinement revealed that disorder can be established as ˜0.9and ˜0.1. This disorder affects the water molecule (O1W); therefore, tokeep the model intact H1W1 was kept with AFIX card (AFIX 1) and thermalparameter at 150% of neighboring O atom.

The HR-XRPD data were collected on D8 Advance diffractometer using CuKα1 radiation (1.54056 Å) with germanium monochromator at RT.Diffraction data were collected in the 2θ range 2-41.5°. Detector scanon solid state LynxEye detector was performed using 0.016° per step with5 sec/step scan speed. The samples were measured in 8 mm long glasscapillary with 0.3 mm outer diameter.

The first step of calculation was taken by finding unit cell parametersusing LSI-Index (Coelho, 2003; Coelho & Kern, 2005) indexing program.The space group was selected on reflections condition and knowledge ofthe compound. The cell parameters, purity as well as instrumentparameters were refined using Whole Powder Pattern Decomposition method(Pawley, 1981).

For Rietveld calculation the cell parameters were taken from the RTmeasurement while the atom positions and their thermal parameters fromsingle crystal measurement file (cif) at 100K. During the refinement thefollowing parameters were refined:

-   -   cell constants;    -   background;    -   instrument geometry;    -   zero shift;    -   absorption.

No atom positions nor thermal motion parameters were refined duringwhole process. The following criteria of fit were used:

-   Y_(o,m) and Y_(c,m) are the observed and calculated data,    respectively at data point m,-   M the number of data points,-   P the number of parameters,-   w_(m) the weighting given to data point m which for counting    statistics is given by wm=1/σ (Y_(o,m)) ² where σ(Y_(o,m)) is the    error in Y_(o,m),

${R_{\exp} = \sqrt{\frac{M - P}{\sum{w_{m}Y_{o,m}^{2}}}}};{R_{wp} = \sqrt{\frac{\sum{w_{m}\left( {T_{o,m} - T_{c,m}} \right)}^{2}}{\sum{w_{m}Y_{o,m}^{2}}}}};{R_{p} = \sqrt{\frac{\sum{{Y_{o,m} - Y_{c,m}}}}{\sum Y_{o,m}}}}$$\mspace{20mu}{{GOF} = {{chi}^{2} = {\frac{R_{wp}}{R_{\exp}} = \sqrt{\frac{\sum{w_{m}\left( {Y_{o,m} - Y_{c,m}} \right)}^{2}}{M - P}}}}}$

The resulting crystal structure is displayed in FIG. 97 and FIG. 98 aswell as a simulation comparison in FIG. 99 which shows close agreementbetween the predicted and experimental diffractograms.

Example 26. Pattern 1 Single Crystal Description

Compound 1 dihydrochloride crystallizes as yellow columnar crystals inthe monoclinic centrosymmetric space group P21/c. In the asymmetricunit, one dicationic API2+, two chloride anions and two water moleculeswere found (overall ratio 1:2:2).

As it can be seen in FIG. 97, the molecule is charged on the most basicamine N atom (N2) as well as on the pyridine N atom (N10). The moleculeadopts an extended (unfolded) conformation.

The crystal is held by intermolecular H-bond interactions (see FIG. 98).One of the chloride anions (ClA) interacts directly with thepositively-charged N atom (N2). The second chloride anion (ClB) acts asa bridge between the two water molecules (see FIG. 97).

However, this ClB interacts also through H-bond with another amine Natom (N14). In the crystal, all water molecules act as donors as well asacceptors of H-bonds, while N atoms act preferably as donors only. Theonly acceptor of H-bond is N16 atom and in this case this interaction isintramolecular.

The intermolecular H-bond network is extended in all three dimensionsand therefore, it produces 3D structure. The HR-XRPD data and Rietveldanalyses performed on starting material delivered by G1 Therapeuticsallowed to obtain cell parameters at RT and revealed that bulk materialconsists of a pure form without any detectable crystalline impurities.Pattern 1 crystallizes in centrosymmetric space group P21/c as adication associated with two chloride anions. Two water molecules werealso found in the asymmetric unit. The results are tabulated below.

TABLE 34 Single crystal data for Pattern 1 Pattern 1 Single CrystalEmpirical formula [(C₂₄H₃₂N₈O)Cl₂•2H₂O] Formula weight 555.51 T [K]100(2) K λ [Å] 0.61993 Å Crystal system Monoclinic Space group P21/cUnit cell dimensions a [Å] 22.118(4) [22.2447(9)] b [Å] 6.9060(14)[6.9472(4)] c [Å] 18.431(4) [18.5906(8)] β [°] 109.58(3) [109.407(2)]V[Å³] 2652.5(10) [2709.7(4)] Z 4 Dc [g/cm³] 1.391 [mm⁻¹] 0.199 F(000)1176 Crystal size [mm³] 0.07 × 0.02 × 0.01 θ range for data collection[°] 1.9-27.6. Reflections collected 45308 Independent reflections 9184[R_(int) = 0.1141] Completeness to θ = 25.242° 99.8 [%] Absorptioncorrection None Max. and min. transmission 0.992 and 0.978Data/restraints/parameters 9183/0/367 Goodness-of-fit on F² 1.019 FinalR indices [I > 2(I)] R1 = 0.0609, wR2 = 0.1271 R indices (all data) R1 =0.1366, wR2 = 0.1603 Absolute configuration Extinction coefficient n/aLargest diff. peak and hole 0.355 and −0.471 [e/Å³]

TABLE 35 Hydrogen bonds from Pattern 1 single crystal structure D-H H .. . A D . . . A D-H . . . A D-H . . . A [Å] [Å] [A] [°] O(1W)—H(1W2) . .. ClA^(i) 1.08 2.20 3.246(3) 162 O(1W)—H(1W1) . . . ClB 1.16(5) 1.92(6)3.064(3) 168(4) O(2W)—H(2W1) . . . 0.86(2) 1.90(2) 2.752(4) 167(2) O(1W)O(2W)—H(2W2) . . . ClB^(ii) 0.88(3) 2.20(3) 3.075(3) 170(2) N(2)—H(2) .. . ClA 0.94(3) 2.13(3) 3.044(2) 163(3) N(10)—H(10) . . . O(2W) 0.95(4)2.08(4) 2.827(3) 135(3) N(10)—H(10) . . . N(16) 0.95(4) 2.05(4) 2.713(3)125(3) N(14)—H(14) . . . ClB^(iii) 0.78(2) 2.45(3) 3.218(2) 171(3)N(23)—H(23) . . . O(22)^(IV) 0.87(3) 1.95(3) 2.814(3) 176(2)

TABLE 36 Final Rietveld parameters for Pattern 1 2theta range (°) 2-41.5R_(exp) 1.94 R_(wp) 3.05 R_(p) 2.36 GOF 1.57 R_(Brag) 1.29 Impurities,other forms [%] below detection limit

Example 27. X-Ray Powder Diffraction (XRPD)

XRPD analysis was carried out on a PANalytical X′pert pro, scanning thefree base samples between 3 and 35° 20. The material was gently groundedto release any agglomerates and loaded onto a multi-well plate withKapton or Mylar polymer film to support the sample. The multi-well platewas then placed into the diffractometer and analyzed using Cu Kradiation (α1 λ=1.54060 Å; α2=1.54443 Å; β=1.39225 Å; α1:α2 ratio=0.5)running in transmission mode (step size 0.0130°2θ) using 40 kV/40 mAgenerator settings.

TABLE 37 XRPD Peak List for Pattern 7. Pos. Height FWHM Left d-spacingRel. Int. [°2θ] [cts] [°2θ] [Å] [%] 5.8473 7100.68 0.0768 15.11477100.00 7.5415 466.35 0.0768 11.72267 6.57 10.8993 2345.84 0.1023 8.1176333.04 11.7793 2076.64 0.1279 7.51310 29.25 14.3833 227.29 0.1023 6.158203.20 15.2147 884.88 0.0895 5.82352 12.46 15.6354 1009.60 0.0895 5.6677514.22 15.9906 516.06 0.1151 5.54265 7.27 16.6298 252.87 0.1023 5.331023.56 17.5834 825.28 0.0895 5.04399 11.62 18.0629 1476.22 0.1279 4.9111620.79 18.2780 645.99 0.0895 4.85384 9.10 18.6556 1724.82 0.1407 4.7564424.29 19.2537 4278.41 0.1151 4.61001 60.25 19.9143 184.16 0.1279 4.458552.59 21.0463 515.45 0.0768 4.22124 7.26 21.6098 3110.68 0.1407 4.1124343.81 22.1211 283.04 0.1023 4.01851 3.99 22.7282 1065.70 0.0895 3.9125415.01 23.3148 2921.40 0.1407 3.81540 41.14 23.6006 520.09 0.0768 3.769847.32 24.0363 142.34 0.1535 3.70249 2.00 24.5873 908.96 0.1535 3.6207512.80 24.8731 374.85 0.1279 3.57978 5.28 25.1286 307.79 0.1023 3.543974.33 27.1701 678.40 0.2047 3.28213 9.55 29.1007 332.84 0.1791 3.068644.69 29.6940 173.72 0.2558 3.00867 2.45 30.2169 163.05 0.1279 2.957792.30 31.4442 135.77 0.2558 2.84508 1.91 33.2805 241.46 0.1791 2.692183.40 33.9789 188.22 0.3070 2.63843 2.65

In certain embodiments, the morphic form is Pattern 7 and ischaracterized by an XRPD pattern comprising at least 2 peaks selectedfrom 5.8±0.2°, 10.9±0.2°, 11.8±0.2°, 15.2±0.2°, 15.6±0.2°, 17.6±0.2°,18.1±0.2°, 18.7±0.2°, 19.3±0.2°, 21.6±0.2°, 22.7±0.2°, 23.3±0.2°, and24.6±0.2°.

In certain embodiments, the morphic form is Pattern 7 and ischaracterized by an XRPD pattern comprising at least 3 peaks selectedfrom 5.8±0.2°, 10.9±0.2°, 11.8±0.2°, 15.2±0.2°, 15.6±0.2°, 17.6±0.2°,18.1±0.2°, 18.7±0.2°, 19.3±0.2°, 21.6±0.2°, 22.7±0.2°, 23.3±0.2°, and24.6±0.2°.

In certain embodiments, the morphic form is Pattern 7 and ischaracterized by an XRPD pattern comprising at least 4 peaks selectedfrom 5.8±0.2°, 10.9±0.2°, 11.8±0.2°, 15.2±0.2°, 15.6±0.2°, 17.6±0.2°,18.1±0.2°, 18.7±0.2°, 19.3±0.2°, 21.6±0.2°, 22.7±0.2°, 23.3±0.2°, and24.6±0.2°.

In certain embodiments, the morphic form is Pattern 7 and ischaracterized by an XRPD pattern comprising at least 5 peaks selectedfrom 5.8±0.2°, 10.9±0.2°, 11.8±0.2°, 15.2±0.2°, 15.6±0.2°, 17.6±0.2°,18.1±0.2°, 18.7±0.2°, 19.3±0.2°, 21.6±0.2°, 22.7±0.2°, 23.3±0.2°, and24.6±0.2°.

In certain embodiments, the morphic form is Pattern 7 and ischaracterized by an XRPD pattern comprising at least 6 peaks selectedfrom 5.8±0.2°, 10.9±0.2°, 11.8±0.2°, 15.2±0.2°, 15.6±0.2°, 17.6±0.2°,18.1±0.2°, 18.7±0.2°, 19.3±0.2°, 21.6±0.2°, 22.7±0.2°, 23.3±0.2°, and24.6±0.2°.

In certain embodiments, the morphic form is Pattern 7 and ischaracterized by an XRPD pattern comprising at least 7 peaks selectedfrom 5.8±0.2°, 10.9±0.2°, 11.8±0.2°, 15.2±0.2°, 15.6±0.2°, 17.6±0.2°,18.1±0.2°, 18.7±0.2°, 19.3±0.2°, 21.6±0.2°, 22.7±0.2°, 23.3±0.2°, and24.6±0.2°.

In certain embodiments, the morphic form is Pattern 7 and ischaracterized by an XRPD pattern comprising at least 8 peaks selectedfrom 5.8±0.2°, 10.9±0.2°, 11.8±0.2°, 15.2±0.2°, 15.6±0.2°, 17.6±0.2°,18.1±0.2°, 18.7±0.2°, 19.3±0.2°, 21.6±0.2°, 22.7±0.2°, 23.3±0.2°, and24.6±0.2°.

In certain embodiments, the morphic form is Pattern 7 and ischaracterized by an XRPD pattern comprising at least 9 peaks selectedfrom 5.8±0.2°, 10.9±0.2°, 11.8±0.2°, 15.2±0.2°, 15.6±0.2°, 17.6±0.2°,18.1±0.2°, 18.7±0.2°, 19.3±0.2°, 21.6±0.2°, 22.7±0.2°, 23.3±0.2°, and24.6±0.2°.

In certain embodiments, the morphic form is Pattern 7 and ischaracterized by an XRPD pattern comprising at least 10 peaks selectedfrom 5.8±0.2°, 10.9±0.2°, 11.8±0.2°, 15.2±0.2°, 15.6±0.2°, 17.6±0.2°,18.1±0.2°, 18.7±0.2°, 19.3±0.2°, 21.6±0.2°, 22.7±0.2°, 23.3±0.2°, and24.6±0.2°.

In certain embodiments, the morphic form is Pattern 7 and ischaracterized by an XRPD pattern comprising at least 11 peaks selectedfrom 5.8±0.2°, 10.9±0.2°, 11.8±0.2°, 15.2±0.2°, 15.6±0.2°, 17.6±0.2°,18.1±0.2°, 18.7±0.2°, 19.3±0.2°, 21.6±0.2°, 22.7±0.2°, 23.3±0.2°, and24.6±0.2°.

In certain embodiments, the morphic form is Pattern 7 and ischaracterized by an XRPD pattern comprising at least 12 peaks selectedfrom 5.8±0.2°, 10.9±0.2°, 11.8±0.2°, 15.2±0.2°, 15.6±0.2°, 17.6±0.2°,18.1±0.2°, 18.7±0.2°, 19.3±0.2°, 21.6±0.2°, 22.7±0.2°, 23.3±0.2°, and24.6±0.2°.

In certain embodiments, the morphic form is Pattern 7 and ischaracterized by an XRPD pattern comprising the 2theta values selectedfrom 5.8±0.2°, 10.9±0.2°, 11.8±0.2°, 18.1±0.2°, 18.7±0.2°, 19.3±0.2°,21.6±0.2°, 23.3±0.2°, and 25.6±0.2°.

TABLE 38 XRPD Peak List for Pattern 8. Pos. Height FWHM Left d-spacingRel. Int. [°2θ] [cts] [°2θ] [Å] [%] 4.0480 347.38 0.0768 21.82815 11.937.4108 547.85 0.0640 11.92907 18.82 8.1013 187.49 0.0512 10.91390 6.449.3731 222.58 0.0640 9.43567 7.65 10.7926 549.61 0.0768 8.19763 18.8815.2279 177.64 0.1023 5.81849 6.10 15.7970 398.88 0.1279 5.61015 13.7016.4382 1158.24 0.0895 5.39272 39.79 17.2971 593.58 0.1023 5.12682 20.3918.4863 2910.63 0.1151 4.79963 100.00 18.7442 1043.18 0.1151 4.7341735.84 19.3171 921.07 0.1407 4.59503 31.64 19.5765 482.48 0.1151 4.5347316.58 20.0240 820.68 0.1023 4.43439 28.20 20.9059 353.65 0.1791 4.2492812.15 21.6070 931.36 0.1023 4.11294 32.00 21.8743 671.04 0.1279 4.0632823.05 23.0044 345.45 0.1023 3.86617 11.87 23.4827 2080.97 0.2047 3.7885071.50 23.8209 1125.13 0.1151 3.73548 38.66 24.1806 1401.67 0.06403.68072 48.16 25.5457 846.89 0.1151 3.48704 29.10 26.3518 179.23 0.12793.38217 6.16 29.5376 209.55 0.2303 3.02424 7.20 29.9908 375.55 0.17912.97957 12.90 31.0626 118.34 0.1535 2.87915 4.07 32.4502 89.31 0.20472.75914 3.07 33.1940 118.10 0.1535 2.69900 4.06 33.9568 101.24 0.15352.64010 3.48

In certain embodiments, the morphic form is Pattern 8 and ischaracterized by an XRPD pattern comprising at least 2 peaks selectedfrom 4.0±0.2°, 7.4±0.2°, 10.8±0.2°, 15.8±0.2°, 16.4±0.2°, 17.3±0.2°,18.5±0.2°, 18.7±0.2°, 19.3±0.2°, 19.6±0.2°, 20.0±0.2°, 20.9±0.2°,21.6±0.2°, 21.9±0.2°, 23.0±0.2°, 23.5±0.2°, 23.8±0.2°, 24.2±0.2°,25.±0.2°, and 29.9±0.2°.

In certain embodiments, the morphic form is Pattern 8 and ischaracterized by an XRPD pattern comprising at least 3 peaks selectedfrom 4.0±0.2°, 7.4±0.2°, 10.8±0.2°, 15.8±0.2°, 16.4±0.2°, 17.3±0.2°,18.5±0.2°, 18.7±0.2°, 19.3±0.2°, 19.6±0.2°, 20.0±0.2°, 20.9±0.2°,21.6±0.2°, 21.9±0.2°, 23.0±0.2°, 23.5±0.2°, 23.8±0.2°, 24.2±0.2°,25.±0.2°, and 29.9±0.2°.

In certain embodiments, the morphic form is Pattern 8 and ischaracterized by an XRPD pattern comprising at least 5 peaks selectedfrom 4.0±0.2°, 7.4±0.2°, 10.8±0.2°, 15.8±0.2°, 16.4±0.2°, 17.3±0.2°,18.5±0.2°, 18.7±0.2°, 19.3±0.2°, 19.6±0.2°, 20.0±0.2°, 20.9±0.2°,21.6±0.2°, 21.9±0.2°, 23.0±0.2°, 23.5±0.2°, 23.8±0.2°, 24.2±0.2°,25.±0.2°, and 29.9±0.2°.

In certain embodiments, the morphic form is Pattern 8 and ischaracterized by an XRPD pattern comprising at least 7 peaks selectedfrom 4.0±0.2°, 7.4±0.2°, 10.8±0.2°, 15.8±0.2°, 16.4±0.2°, 17.3±0.2°,18.5±0.2°, 18.7±0.2°, 19.3±0.2°, 19.6±0.2°, 20.0±0.2°, 20.9±0.2°,21.6±0.2°, 21.9±0.2°, 23.0±0.2°, 23.5±0.2°, 23.8±0.2°, 24.2±0.2°,25.±0.2°, and 29.9±0.2°.

In certain embodiments, the morphic form is Pattern 8 and ischaracterized by an XRPD pattern comprising at least 9 peaks selectedfrom 4.0±0.2°, 7.4±0.2°, 10.8±0.2°, 15.8±0.2°, 16.4±0.2°, 17.3±0.2°,18.5±0.2°, 18.7±0.2°, 19.3±0.2°, 19.6±0.2°, 20.0±0.2°, 20.9±0.2°,21.6±0.2°, 21.9±0.2°, 23.0±0.2°, 23.5±0.2°, 23.8±0.2°, 24.2±0.2°,25.±0.2°, and 29.9±0.2°.

In certain embodiments, the morphic form is Pattern 8 and ischaracterized by an XRPD pattern comprising at least 11 peaks selectedfrom 4.0±0.2°, 7.4±0.2°, 10.8±0.2°, 15.8±0.220 , 16.4±0.2°, 17.3±0.2°,18.5±0.2°, 18.7±0.2°, 19.3±0.2°, 19.6±0.2°, 20.0±0.2°, 20.9±0.2°,21.6±0.2°, 21.9±0.2°, 23.0±0.2°, 23.5±0.2°, 23.8±0.2°, 24.2±0.2°,25.±0.2°, and 29.9±0.2°.

In certain embodiments, the morphic form is Pattern 8 and ischaracterized by an XRPD pattern comprising at least 13 peaks selectedfrom 4.0±0.2°, 7.4±0.2°, 10.8±0.2°, 15.8±0.2°, 16.4±0.2°, 17.3±0.2°,18.5±0.2°, 18.7±0.2°, 19.3±0.2°, 19.6±0.2°, 20.0±0.2°, 20.9±0.2°,21.6±0.2°, 21.9±0.2°, 23.0±0.2°, 23.5±0.2°, 23.8±0.2°, 24.2±0.2°,25.±0.2°, and 29.9±0.2°.

In certain embodiments, the morphic form is Pattern 8 and ischaracterized by an XRPD pattern comprising at least 15 peaks selectedfrom 4.0±0.2°, 7.4±0.2°, 10.8±0.2°, 15.8±0.2°, 16.4±0.2°, 17.3±0.2°,18.5±0.2°, 18.7±0.2°, 19.3±0.2°, 19.6±0.2°, 20.0±0.2°, 20.9±0.2°,21.6±0.2°, 21.9±0.2°, 23.0±0.2°, 23.5±0.2°, 23.8±0.2°, 24.2±0.2°,25.±0.2°, and 29.9±0.2°.

In certain embodiments, the morphic form is Pattern 8 and ischaracterized by an XRPD pattern comprising at least 17 peaks selectedfrom 4.0±0.2°, 7.4±0.2°, 10.8±0.2°, 15.8±0.2°, 16.4±0.2°, 17.3±0.2°,18.5±0.2°, 18.7±0.2°, 19.3±0.2°, 19.6±0.2°, 20.0±0.2°, 20.9±0.2°,21.6±0.2°, 21.9±0.2°, 23.0±0.2°, 23.5±0.2°, 23.8±0.2°, 24.2±0.2°,25.±0.2°, and 29.9±0.2°.

In certain embodiments, the morphic form is Pattern 8 and ischaracterized by an XRPD pattern comprising at least 19 peaks selectedfrom 4.0±0.2°, 7.4±0.2°, 10.8±0.2°, 15.8±0.2°, 16.4±0.2°, 17.3±0.2°,18.5±0.2°, 18.7±0.2°, 19.3±0.2°, 19.6±0.2°, 20.0±0.2°, 20.9±0.2°,21.6±0.2°, 21.9±0.2°, 23.0±0.2°, 23.5±0.2°, 23.8±0.2°, 24.2±0.2°,25.±0.2°, and 29.9±0.2°.

In certain embodiments, the morphic form is Pattern 8 and ischaracterized by an XRPD pattern comprising the 2theta values selectedfrom 4.0±0.2°, 7.4±0.2°, 10.8±0.2°, 15.8±0.2°, 16.4±0.2°, 17.3±0.2°,18.5±0.2°, 18.7±0.2°, 19.3±0.2°, 19.6±0.2°, 20.0±0.2°, 20.9±0.2°,21.6±0.2°, 21.9±0.2°, 23.0±0.2°, 23.5±0.2°, 23.8±0.2°, 24.2±0.2°,25.±0.2°, and 29.9±0.2°.

TABLE 39 XRPD Peak List for Pattern 9. Pos. Height FWHM Left d-spacingRel. Int. [°2θ] [cts] [°2θ] [Å] [%] 3.3241 66.84 0.6140 26.58015 1.225.8249 435.60 0.0768 15.17296 7.95 10.8757 163.63 0.0768 8.13513 2.9811.2250 455.75 0.0384 7.88278 8.31 11.3768 2582.29 0.0256 7.77792 47.1011.4550 3239.65 0.0512 7.72505 59.09 11.7506 125.87 0.1023 7.53135 2.3012.7555 911.61 0.0768 6.94019 16.63 16.2943 579.18 0.0640 5.44001 10.5616.9724 5482.56 0.0640 5.22416 100.00 18.3347 317.87 0.0768 4.83897 5.8019.0290 1955.36 0.0512 4.66393 35.67 21.6233 209.25 0.1023 4.10989 3.8222.5372 3327.96 0.0640 3.94525 60.70 23.0204 228.23 0.0895 3.86352 4.1623.3310 202.17 0.1023 3.81279 3.69 24.0081 1586.58 0.0768 3.70676 28.9424.7459 1138.05 0.0512 3.59790 20.76 25.0321 671.51 0.0512 3.55741 12.2525.1807 628.82 0.0384 3.53675 11.47 25.6410 589.48 0.0512 3.47430 10.7526.8890 774.68 0.0384 3.31581 14.13 27.3292 336.97 0.1535 3.26339 6.1528.1945 1400.08 0.0768 3.16518 25.54 29.2218 587.48 0.0768 3.05620 10.7229.5880 578.30 0.0468 3.01670 10.55 29.6825 2105.96 0.0624 3.00731 38.4129.7556 2789.47 0.0512 3.00258 50.88 30.8547 227.32 0.0768 2.89808 4.1531.3680 103.61 0.1791 2.85182 1.89 32.5428 1281.07 0.0624 2.74922 23.3732.6311 1246.75 0.0468 2.74880 22.74 33.0463 389.09 0.0468 2.70848 7.1033.1273 426.22 0.0468 2.70204 7.77 33.6270 830.91 0.0624 2.66302 15.1634.2293 196.40 0.1248 2.61753 3.58 34.4200 366.04 0.0468 2.60346 6.6834.5206 224.05 0.0468 2.60256 4.09

In certain embodiments, the morphic form is Pattern 9 and ischaracterized by an XRPD pattern comprising at least 2 peaks selectedfrom 11.4±0.2°, 11.5±0.2°, 12.8±0.2°, 16.3±0.2°, 16.9±0.2°, 19.0±0.2°,22.5±0.2°, 24.0±0.2°, 24.7±0.2°, 25.0±0.2°, 25.2±0.2°, 25.6±0.2°,26.9±0.2°, 28.2±0.2°, 29.2±0.2°, 29.6±0.2°, 29.7±0.2°, 29.8±0.2°,32.5±0.2°, 32.6±0.2°, and 33.6±0.2°.

In certain embodiments, the morphic form is Pattern 9 and ischaracterized by an XRPD pattern comprising at least 3 peaks selectedfrom 11.4±0.2°, 11.5±0.2°, 12.8±0.2°, 16.3±0.2°, 16.9±0.2°, 19.0±0.2°,22.5±0.2°, 24.0±0.2°, 24.7±0.2°, 25.0±0.2°, 25.2±0.2°, 25.6±0.2°,26.9±0.2°, 28.2±0.2°, 29.2±0.2°, 29.6±0.2°, 29.7±0.2°, 29.8±0.2°,32.5±0.2°, 32.6±0.2°, and 33.6±0.2°.

In certain embodiments, the morphic form is Pattern 9 and ischaracterized by an XRPD pattern comprising at least 5 peaks selectedfrom 11.4±0.2°, 11.5±0.2°, 12.8±0.2°, 16.3±0.2°, 16.9±0.2°, 19.0±0.2°,22.5±0.2°, 24.0±0.2°, 24.7±0.2°, 25.0±0.2°, 25.2±0.2°, 25.6±0.2°,26.9±0.2°, 28.2±0.2°, 29.2±0.2°, 29.6±0.2°, 29.7±0.2°, 29.8±0.2°,32.5±0.2°, 32.6±0.2°, and 33.6±0.2°.

In certain embodiments, the morphic form is Pattern 9 and ischaracterized by an XRPD pattern comprising at least 7 peaks selectedfrom 11.4±0.2°, 11.5±0.2°, 12.8±0.2°, 16.3±0.2°, 16.9±0.2°, 19.0±0.2°,22.5±0.2°, 24.0±0.2°, 24.7±0.2°, 25.0±0.2°, 25.2±0.2°, 25.6±0.2°,26.9±0.2°, 28.2±0.2°, 29.2±0.2°, 29.6±0.2°, 29.7±0.2°, 29.8±0.2°,32.5±0.2°, 32.6±0.2°, and 33.6±0.2°.

In certain embodiments, the morphic form is Pattern 9 and ischaracterized by an XRPD pattern comprising at least 9 peaks selectedfrom 11.4±0.2°, 11.5±0.2°, 12.8±0.2°, 16.3±0.2°, 16.9±0.2°, 19.0±0.2°,22.5±0.2°, 24.0±0.2°, 24.7±0.2°, 25.0±0.2°, 25.2±0.2°, 25.6±0.2°,26.9±0.2°, 28.2±0.2°, 29.2±0.2°, 29.6±0.2°, 29.7±0.2°, 29.8±0.2°,32.5±0.2°, 32.6±0.2°, and 33.6±0.2°.

In certain embodiments, the morphic form is Pattern 9 and ischaracterized by an XRPD pattern comprising at least 11 peaks selectedfrom 11.4±0.2°, 11.5±0.2°, 12.8±0.2°, 16.3±0.2°, 16.9±0.2°, 19.0±0.2°,22.5±0.2°, 24.0±0.2°, 24.7±0.2°, 25.0±0.2°, 25.2±0.2°, 25.6±0.2°,26.9±0.2°, 28.2±0.2°, 29.2±0.2°, 29.6±0.2°, 29.7±0.2°, 29.8±0.2°,32.5±0.2°, 32.6±0.2°, and 33.6±0.2°.

In certain embodiments, the morphic form is Pattern 9 and ischaracterized by an XRPD pattern comprising at least 13 peaks selectedfrom 11.4±0.2°, 11.5±0.2°, 12.8±0.2°, 16.3±0.2°, 16.9±0.2°, 19.0±0.2°,22.5±0.2°, 24.0±0.2°, 24.7±0.2°, 25.0±0.2°, 25.2±0.2°, 25.6±0.2°,26.9±0.2°, 28.2±0.2°, 29.2±0.2°, 29.6±0.2°, 29.7±0.2°, 29.8±0.2°,32.5±0.2°, 32.6±0.2°, and 33.6±0.2°.

In certain embodiments, the morphic form is Pattern 9 and ischaracterized by an XRPD pattern comprising at least 15 peaks selectedfrom 11.4±0.2°, 11.5±0.2°, 12.8±0.2°, 16.3±0.2°, 16.9±0.2°, 19.0±0.2°,22.5±0.2°, 24.0±0.2°, 24.7±0.2°, 25.0±0.2°, 25.2±0.2°, 25.6±0.2°,26.9±0.2°, 28.2±0.2°, 29.2±0.2°, 29.6±0.2°, 29.7±0.2°, 29.8±0.2°,32.5±0.2°, 32.6±0.2°, and 33.6±0.2°.

In certain embodiments, the morphic form is Pattern 9 and ischaracterized by an XRPD pattern comprising at least 17 peaks selectedfrom 11.4±0.2°, 11.5±0.2°, 12.8±0.2°, 16.3±0.2°, 16.9±0.2°, 19.0±0.2°,22.5±0.2°, 24.0±0.2°, 24.7±0.2°, 25.0±0.2°, 25.2±0.2°, 25.6±0.2°,26.9±0.2°, 28.2±0.2°, 29.2±0.2°, 29.6±0.2°, 29.7±0.2°, 29.8±0.2°,32.5±0.2°, 32.6±0.2°, and 33.6±0.2°.

In certain embodiments, the morphic form is Pattern 9 and ischaracterized by an XRPD pattern comprising at least 19 peaks selectedfrom 11.4±0.2°, 11.5±0.2°, 12.8±0.2°, 16.3±0.2°, 16.9±0.2°, 19.0±0.2°,22.5±0.2°, 24.0±0.2°, 24.7±0.2°, 25.0±0.2°, 25.2±0.2°, 25.6±0.2°,26.9±0.2°, 28.2±0.2°, 29.2±0.2°, 29.6±0.2°, 29.7±0.2°, 29.8±0.2°,32.5±0.2°, 32.6±0.2°, and 33.6±0.2°.

In certain embodiments, the morphic form is Pattern 9 and ischaracterized by an XRPD pattern comprising the 2theta values selectedfrom 11.4±0.2°, 11.5±0.2°, 12.8±0.2°, 16.3±0.2°, 16.9±0.2°, 19.0±0.2°,22.5±0.2°, 24.0±0.2°, 24.7±0.2°, 25.0±0.2°, 25.2±0.2°, 25.6±0.2°,26.9±0.2°, 28.2±0.2°, 29.2±0.2°, 29.6±0.2°, 29.7±0.2°, 29.8±0.2°,32.5±0.2°, 32.6±0.2°, and 33.6±0.2°.

TABLE 40 XRPD Peak List for Pattern 10. Pos. Height FWHM Left d-spacingRel. Int. [°2θ] [cts] [°2θ] [Å] [%] 5.8105 111.76 0.1023 15.21055 3.028.4576 348.24 0.0512 10.45487 9.42 8.6052 938.19 0.0384 10.27586 25.378.8323 3698.48 0.0895 10.01221 100.00 10.8959 47.29 0.1535 8.12011 1.2811.7796 49.97 0.2047 7.51285 1.35 17.7257 115.24 0.2047 5.00383 3.1218.7729 461.23 0.0768 4.72700 12.47 19.2217 201.82 0.1279 4.61762 5.4619.8311 260.56 0.1023 4.47708 7.04 21.6431 180.47 0.1279 4.10617 4.8822.7827 1974.03 0.1023 3.90330 53.37 23.3096 166.09 0.1535 3.81624 4.4924.3184 133.94 0.1279 3.66017 3.62 26.3881 331.12 0.1023 3.37760 8.9526.7437 104.59 0.1535 3.33350 2.83 27.6105 180.69 0.1535 3.23079 4.8928.3607 1012.39 0.0895 3.14701 27.37 30.0318 3097.78 0.1279 2.9755983.76 30.4202 255.52 0.0768 2.93848 6.91 30.6379 1479.82 0.0468 2.9156840.01 30.7217 2244.78 0.0512 2.91033 60.69 31.1380 428.31 0.0895 2.8723611.58 31.3221 416.14 0.1279 2.85589 11.25 31.9484 263.81 0.0512 2.801327.13 32.4521 119.39 0.1279 2.75899 3.23 33.1508 177.25 0.0768 2.702414.79 33.4324 1032.02 0.1151 2.68030 27.90 34.3039 136.16 0.1535 2.614173.68 34.6150 319.87 0.2047 2.59138 8.65

In certain embodiments, the morphic form is Pattern 10 and ischaracterized by an XRPD pattern comprising at least 2 peaks selectedfrom 8.6±0.2°, 8.8±0.2°, 18.8±0.2°, 22.8±0.2°, 28.4±0.2°, 30.0±0.2°,30.6±0.2°, 30.7±0.2°, 31.1±0.2°, 31.3±0.2°, and 33.4±0.2°.

In certain embodiments, the morphic form is Pattern 10 and ischaracterized by an XRPD pattern comprising at least 3 peaks selectedfrom 8.6±0.2°, 8.8±0.2°, 18.8±0.2°, 22.8±0.2°, 28.4±0.2°, 30.0±0.2°,30.6±0.2°, 30.7±0.2°, 31.1±0.2°, 31.3±0.2°, and 33.4±0.2°.

In certain embodiments, the morphic form is Pattern 10 and ischaracterized by an XRPD pattern comprising at least 5 peaks selectedfrom 8.6±0.2°, 8.8±0.2°, 18.8±0.2°, 22.8±0.2°, 28.4±0.2°, 30.0±0.2°,30.6±0.2°, 30.7±0.2°, 31.1±0.2°, 31.3±0.2°, and 33.4±0.2°.

In certain embodiments, the morphic form is Pattern 10 and ischaracterized by an XRPD pattern comprising at least 7 peaks selectedfrom 8.6±0.2°, 8.8±0.2°, 18.8±0.2°, 22.8±0.2°, 28.4±0.2°, 30.0±0.2°,30.6±0.2°, 30.7±0.2°, 31.1±0.2°, 31.3±0.2°, and 33.4±0.2°.

In certain embodiments, the morphic form is Pattern 10 and ischaracterized by an XRPD pattern comprising at least 9 peaks selectedfrom 8.6±0.2°, 8.8±0.2°, 18.8±0.2°, 22.8±0.2°, 28.4±0.2°, 30.0±0.2°,30.6±0.2°, 30.7±0.2°, 31.1±0.2°, 31.3±0.2°, and 33.4±0.2°.

In certain embodiments, the morphic form is Pattern 10 and ischaracterized by an XRPD pattern comprising the 2theta values selectedfrom 8.6±0.2°, 8.8±0.2°, 18.8±0.2°, 22.8±0.2°, 28.4±0.2°, 30.0±0.2°,30.6±0.2°, 30.7±0.2°, 31.1±0.2°, 31.3±0.2°, and 33.4±0.2°.

This specification has been described with reference to embodiments ofthe invention. However, one of ordinary sill in the art appreciates thatvarious modifications and changes can be made without departing from thescope of the invention as set forth in the claims below. Accordingly,the specification is to be regarded in an illustrative rather than arestrictive sense, and all such modifications are intended to beincluded within the scope of invention.

We claim:
 1. A crystalline compound of structure:

which is a dihydrochloride.
 2. The crystalline compound of claim 1characterized by an X-ray powder diffraction (XRPD) pattern comprisingat least three 2theta values selected from 9.6±0.2°, 21.3±0.2°,19.8±0.2°, 12.2±0.2°, 24.0±0.2°, 26.1±0.2°, 19.3±0.2°, 17.6±0.2°, and28.6±0.2°.
 3. The crystalline compound of claim 2, wherein the XRPDpattern comprises at least four 2theta values selected from 9.6±0.2°,21.3±0.2°, 19.8±0.2°, 12.2±0.2°, 24.0±0.2°, 26.1±0.2°, 19.3±0.2°,17.6±0.2°, and 28.6±0.2°.
 4. The crystalline compound of claim 2,wherein the XRPD pattern comprises at least five 2theta values selectedfrom 9.6±0.2°, 21.3±0.2°, 19.8±0.2°, 12.2±0.2°, 24.0±0.2°, 26.1±0.2°,19.3±0.2°, 17.6±0.2°, and 28.6±0.2°.
 5. The crystalline compound ofclaim 2, wherein the XRPD pattern comprises at least six 2theta valuesselected from 9.6±0.2°, 21.3±0.2°, 19.8±0.2°, 12.2±0.2°, 24.0±0.2°,26.1±0.2°, 19.3±0.2°, 17.6±0.2°, and 28.6±0.2°.
 6. The crystallinecompound of claim 2, wherein the XRPD pattern comprises at least the2theta value of 9.6±0.2°.
 7. The crystalline compound of claim 2,wherein the XRPD pattern comprises at least the 2theta values of9.6±0.2°, 19.8±0.2°, and 21.3±0.2°.
 8. A composition comprising thecrystalline compound of claim 1, mannitol and citric acid.
 9. Apharmaceutical composition comprising the crystalline compound of claim1 and a pharmaceutically acceptable excipient.
 10. The pharmaceuticalcomposition of claim 9, wherein the pharmaceutically acceptableexcipient comprises mannitol.
 11. The pharmaceutical composition ofclaim 9, wherein the pharmaceutically acceptable excipient comprisescitric acid.
 12. The pharmaceutical composition of claim 9, wherein thepharmaceutically acceptable excipient comprises mannitol and citricacid.
 13. The pharmaceutical composition of claim 9 comprising about200-600 milligrams of the crystalline compound.
 14. The pharmaceuticalcomposition of claim 9 comprising about 300 milligrams of thecrystalline compound.
 15. The pharmaceutical composition of claim 9comprising about 349 milligrams of the crystalline compound.
 16. Thepharmaceutical composition of claim 10 comprising about 250-350 mg ofmannitol.
 17. The pharmaceutical composition of claim 10 comprisingabout 300 mg of mannitol.
 18. The pharmaceutical composition of claim 11comprising about 50-100 mg of citric acid.
 19. The pharmaceuticalcomposition of claim 11 comprising about 76 mg of citric acid.
 20. Thepharmaceutical composition of claim 12 comprising about 250-350 mg ofmannitol.
 21. The pharmaceutical composition of claim 12 comprisingabout 300 mg of mannitol.
 22. The pharmaceutical composition of claim 12comprising about 50-100 mg of citric acid.
 23. The pharmaceuticalcomposition of claim 12 comprising about 76 mg of citric acid.
 24. Thepharmaceutical composition of claim 12 comprising about 300 mg ofmannitol and about 76 mg of citric acid.
 25. The pharmaceuticalcomposition of claim 24 comprising about 300 milligrams of thecrystalline compound.
 26. The pharmaceutical composition of claim 12comprising a dose of about 150 mg/m² to about 350 mg/m² of thecrystalline compound.
 27. The pharmaceutical composition of claim 12comprising a dose of about 240 mg/m² of the crystalline compound.